Pivotal role of xanthine oxidase in the initiation of tubulointerstitial renal injury in rats with hyperlipidemia
2006; Elsevier BV; Volume: 69; Issue: 3 Linguagem: Inglês
10.1038/sj.ki.5000121
ISSN1523-1755
AutoresWilfried Gwinner, Hartmut Scheuer, Hermann Haller, Ralf P. Brandes, Hermann-Josef Groene,
Tópico(s)Dialysis and Renal Disease Management
ResumoHyperlipidemia can induce or aggravate renal tubulointerstitial injury. Experiments in a complex rat model with chronic glomerulonephritis and long-standing, coexisting hyperlipidemia suggested that induction of xanthine oxidase (XO), with increased oxygen radical generation, is involved in aggravation of tubulointerstitial injury. To separate the role of XO in the initial events of lipid-mediated tubulointerstitial injury, short-term experiments with diet-induced hyperlipidemia over 21 and 35 days were performed in otherwise healthy rats. XO expression in relation to the antioxidant enzymes was examined in the cortical tubulointerstitium (TIS) and proximal tubules (PT). Subsequent experiments with XO inhibition were performed, examining tubulointerstitial infiltration with ED1-positive cells and expression of adhesion molecules and monocyte chemoattractant protein-1 (MCP-1) as indicators of early injurious events. Hyperlipidemia increased XO activity in TIS by 40 and 86%, and in PT by 28 and 90% at days 21 and 35, compared with controls on regular diet. This increased activity was associated with increased reactive oxygen species. Among the antioxidant enzymes, glutathione peroxidase activity increased in TIS by 40% and in PT by 90%. Histological evaluation showed a three-fold increase in ED1-positive cells and increased MCP-1 and vascular cell adhesion molecule-1 (VCAM-1) expression at day 35 in the TIS. Inhibition of XO prevented tubulointerstitial ED1 cell infiltration, together with a decreased expression of MCP-1 and VCAM-1. These results point to an important role for XO in the early stage of hyperlipidemia-associated renal injury, mediating macrophage infiltration by a putatively redox-dependent upregulation of MCP-1 and VCAM-1. Hyperlipidemia can induce or aggravate renal tubulointerstitial injury. Experiments in a complex rat model with chronic glomerulonephritis and long-standing, coexisting hyperlipidemia suggested that induction of xanthine oxidase (XO), with increased oxygen radical generation, is involved in aggravation of tubulointerstitial injury. To separate the role of XO in the initial events of lipid-mediated tubulointerstitial injury, short-term experiments with diet-induced hyperlipidemia over 21 and 35 days were performed in otherwise healthy rats. XO expression in relation to the antioxidant enzymes was examined in the cortical tubulointerstitium (TIS) and proximal tubules (PT). Subsequent experiments with XO inhibition were performed, examining tubulointerstitial infiltration with ED1-positive cells and expression of adhesion molecules and monocyte chemoattractant protein-1 (MCP-1) as indicators of early injurious events. Hyperlipidemia increased XO activity in TIS by 40 and 86%, and in PT by 28 and 90% at days 21 and 35, compared with controls on regular diet. This increased activity was associated with increased reactive oxygen species. Among the antioxidant enzymes, glutathione peroxidase activity increased in TIS by 40% and in PT by 90%. Histological evaluation showed a three-fold increase in ED1-positive cells and increased MCP-1 and vascular cell adhesion molecule-1 (VCAM-1) expression at day 35 in the TIS. Inhibition of XO prevented tubulointerstitial ED1 cell infiltration, together with a decreased expression of MCP-1 and VCAM-1. These results point to an important role for XO in the early stage of hyperlipidemia-associated renal injury, mediating macrophage infiltration by a putatively redox-dependent upregulation of MCP-1 and VCAM-1. Hyperlipidemia is common in patients with chronic renal failure. Some clinical studies have suggested that hyperlipidemia in patients with renal disease may act as a modulating factor that contributes to progressive renal impairment.1.Attman P.O. Alaupovic P. Samuelsson O. Lipoprotein abnormalities as a risk factor for progressive nondiabetic renal disease.Kidney Int. 1999; 56: S14-S17Abstract Full Text Full Text PDF Google Scholar, 2.Syrjanen J. Mustonen J. Pasternack A. Hypertriglyceridaemia and hyperuricaemia are risk factors for progression of IgA nephropathy.Nephrol Dial Transplant. 2000; 15: 34-42Crossref PubMed Scopus (234) Google Scholar, 3.Massy Z.A. Khoa T.N. Lacour B. et al.Dyslipidaemia and the progression of renal disease in chronic renal failure patients.Nephrol Dial Transplant. 1999; 14: 2392-2397Crossref PubMed Scopus (66) Google Scholar, 4.Samuelsson O. Mulec H. Knight-Gibson C. et al.Lipoprotein abnormalities are associated with increased rate of progression of human chronic renal insufficiency.Nephrol Dial Transplant. 1997; 12: 1908-1915Crossref PubMed Scopus (215) Google Scholar This is supported by diverse experimental studies showing increased glomerular and tubulointerstitial damage and macrophage infiltration, higher protein excretion, and accelerated loss of renal function in hyperlipidemic animals.5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 6.Grone H.J. Walli A. Grone E. et al.Induction of glomerulosclerosis by dietary lipids. A functional and morphologic study in the rat.Lab Invest. 1989; 60: 433-446PubMed Google Scholar, 7.Kasiske B.L. O'Donnell M.P. Cleary M.P. Keane W.F. Effects of reduced renal mass on tissue lipids and renal injury in hyperlipidemic rats.Kidney Int. 1989; 35: 40-47Abstract Full Text PDF PubMed Scopus (63) Google Scholar Demonstration of oxidatively modified lipids in the kidney and renal protective effects with vitamin E supplementation suggested that oxidative injury is involved in the mechanisms of lipid-mediated tissue damage.5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 8.Lee H.S. Jeong J.Y. Kim B.C. et al.Dietary antioxidant inhibits lipoprotein oxidation and renal injury in experimental focal segmental glomerulosclerosis.Kidney Int. 1997; 51: 1151-1159Abstract Full Text PDF PubMed Scopus (83) Google Scholar We have recently examined the long-term effects of hyperlipidemia in rats with chronic anti-Thy1 glomerulonephritis.9.Scheuer H. Gwinner W. Hohbach J. et al.Oxidant stress in hyperlipidemia-induced renal damage.Am J Physiol. 2000; 278: F63-F74PubMed Google Scholar Compared with nephritic animals without hyperlipidemia, animals with nephritis and hyperlipidemia over a period of 70 and 150 days showed accelerated glomerulosclerosis and tubulointerstitial damage, including tubular atrophy and tubulointerstitial macrophage infiltration and fibrosis. Accelerated renal injury was associated with an increased generation of reactive oxygen species (ROS) and a decrease in intrinsic antioxidant enzyme activities in the tubulointerstitium. Moreover, oxidatively modified proteins were detected in renal tissue, indicative of oxidative stress and injury. As the major cause of increased ROS generation, we identified xanthine oxidase. The activity of this oxidase was two- to three-fold higher in the tubulointerstitium, together with increased quantities of the enzyme as determined by Western blot analysis. Based on these results, xanthine oxidase-dependent oxidative pathogenetic mechanisms could play an important role in hyperlipidemia-mediated renal damage. However, as our previous studies were performed in a complex model with uninephrectomy and glomerulonephritis and at an advanced stage of renal injury, we could not determine whether xanthine oxidase is involved in the initial sequence of changes in the tubulointerstitium caused by hyperlipidemia. Therefore, the present study examines the isolated and early effect of hyperlipidemia on xanthine oxidase expression and tubulointerstitial changes in rats without other coexisting renal pathologies. The high-fat and high-cholesterol diet resulted in a significant increase in cholesterol and triglyceride serum levels. Serum creatinine levels and urinary protein excretion were unchanged in hyperlipidemic animals. Tubular atrophy and tubulointerstitial fibrosis were absent in controls and hyperlipidemic animals (not shown). Staining for ED1-positive cells revealed a moderate increase in infiltrating cells in the cortical tubulointerstitium of the animals with hyperlipidemia. Taken together, these minor changes indicated an early stage of lipid-mediated renal alterations.Table 1Laboratory and immunohistochemistry data in experimental animalsControlsHLP 21 daysHLP 35 daysCholesterol in serum (mg/dl)71±9107±32**107±19**(n=17)(n=8)(n=8)Triglycerides in serum (mg/dl)36±1881±35*60±17*(n=16)(n=8)(n=8)Serum creatinine (mg/dl)0.50±0.040.52±0.020.55±0.05(n=17)(n=6)(n=7)Protein excretion (mg/24 h)19.3±7.318.3±6.215.3±5.2(n=17)(n=8)(n=8)ED1-positive cells3.2±0.86.1±2.3**9.9±6.2**(number/high-power field)aMagnification: × 400.(n=17)(n=8)(n=8)Controls: animals with standard diet. HLP 21 days and HLP 35 days: hyperlipidemic animals on a fat- and cholesterol-enriched diet for 21 or 35 days, respectively. Values are means±s.d.; *P<0.05 vs controls; **P<0.01 vs controls.a Magnification: × 400. Open table in a new tab Controls: animals with standard diet. HLP 21 days and HLP 35 days: hyperlipidemic animals on a fat- and cholesterol-enriched diet for 21 or 35 days, respectively. Values are means±s.d.; *P<0.05 vs controls; **P<0.01 vs controls. Chemiluminescence (CL) measurements in tubulointerstitial samples and in specifically isolated proximal tubules with luminol, a general detector of ROS, indicated an increased generation of ROS in both tissue fractions of hyperlipidemic animals (Figure 1a). CL with lucigenin was increased in the tubulointerstitial fraction at day 35, indicating an involvement of superoxide anion radical (Figure 1b). Increased ROS generation was associated with higher activities of xanthine oxidase in the tubulointerstitium and in proximal tubules (Figure 2). Further evaluation of xanthine oxidase expression in proximal tubules by Western and Northern blotting revealed increased protein and mRNA levels of the enzyme in hyperlipidemic animals (Figure 3). By immunohistochemistry, xanthine oxidase expression was localized in the peritubular area, most probably corresponding to peritubular capillaries in controls and in hyperlipidemic animals. In addition, animals with hyperlipidemia showed staining of xanthine oxidase in tubular epithelial cells and in the vicinity of mononuclear cells infiltrating the region of peritubular capillaries (Figure 4).Figure 3Levels of xanthine oxidoreductase (XOR) mRNA and protein in proximal tubules of control animals with standard diet and hyperlipidemic animals (HLP) fed a fat- and cholesterol-enriched diet for 35 days. Results are representative of independent analyses of four samples from the controls and six samples from hyperlipidemic animals. (a) Levels of XOR mRNA are shown in relation to the 28S RNA of the samples. Relative optical densities for XOR mRNA after correction for 28S RNA abundance were 1.0±0.3 in the control samples and 1.4±0.2 in the samples from animals with hyperlipidemia. (b) XOR protein bands are shown in relation to the internal loading control β-tubulin. Splitted protein bands correspond to the two major fragments of XOR, which are present under reducing conditions.10.Sarnesto A. Linder N. Raivio K.O. Organ distribution and molecular forms of human xanthine dehydrogenase/xanthine oxidase protein.Lab Invest. 1996; 74: 48-56PubMed Google Scholar Relative optical densities for XOR after correction for β-tubulin abundance were 1.9±0.4 in the control samples and 3.0±0.5 in the samples from animals with hyperlipidemia. Right lane: molecular weight marker proteins.View Large Image Figure ViewerDownload (PPT)Figure 4Immunohistology for xanthine oxidase in the cortical tubulointerstitium. (a) In the controls without hyperlipidemia, xanthine oxidase staining is present in the peritubular region. (b) In animals with hyperlipidemia, increased xanthine oxidase expression is detected in the tubular epithelium, in the peritubular area probably corresponding to peritubular capillaries, and in the vicinity of mononuclear cells (arrows). Original magnification × 400.View Large Image Figure ViewerDownload (PPT) Despite increased ROS generation, there were only minor changes in the expression of antioxidant enzymes in tubulointerstitial and proximal tubular samples of hyperlipidemic animals (Figure 5). Total activities of superoxide dismutase (SOD) (i.e. copper/zinc SOD and manganese SOD), manganese SOD activities alone, as well as catalase activities remained virtually unchanged. Of note, glutathione peroxidase showed an increased activity in hyperlipidemic animals that was maximal in proximal tubules after 35 days of high-fat and high-cholesterol diet. Based on the putative role of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and monocyte chemoattractant protein-1 (MCP-1) in the tubulointerstitial infiltration with macrophages in hyperlipidemia5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 11.Hattori M. Nikolic-Paterson D.J. Miyazaki K. et al.Mechanisms of glomerular macrophage infiltration in lipid-induced renal injury.Kidney Int. 1999; 56: S47-S50Abstract Full Text Full Text PDF Google Scholar, 12.Kodama N. Otani H. Yamada Y. et al.Involvement of MCP-1 and M-CSF in glomerular foam cell formation in ExHC rats.Kidney Int. 1999; 56: S174-S177Abstract Full Text PDF Google Scholar and the reported ROS-dependent regulation of these factors,13.Marui N. Offermann M.K. Swerlick R. et al.Vascular cell adhesion molecule-1 (VCAM-1) gene transcription and expression are regulated through an antioxidant-sensitive mechanism in human vascular endothelial cells.J Clin Invest. 1993; 92: 1866-1874Crossref PubMed Scopus (954) Google Scholar, 14.Lakshminarayanan V. Beno D.W. Costa R.H. Roebuck K.A. Differential regulation of interleukin-8 and intercellular adhesion molecule-1 by H2O2 and tumor necrosis factor-alpha in endothelial and epithelial cells.J Biol Chem. 1997; 272: 32910-32918Crossref PubMed Scopus (141) Google Scholar, 15.Satriano J.A. Shuldiner M. Hora K. et al.Oxygen radicals as second messengers for expression of the monocyte chemoattractant protein JE/MCP-1, and the monocyte colony-stimulating factor, CSF-1, in response to tumor necrosis factor-alpha and immunoglobulin G. Evidence for involvement of reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent oxidase.J Clin Invest. 1993; 92: 1564-1571Crossref PubMed Scopus (279) Google Scholar we hypothesized that xanthine oxidase might be involved in the process of macrophage infiltration in our model. Therefore, experiments were performed with animals in which xanthine oxidase had been inhibited by administration of tungsten. Tungsten treatment did not significantly change cholesterol and triglyceride serum levels in animals fed a high-cholesterol and high-fat diet (cholesterol: 95±20 vs 94±24 mg/dl, triglycerides 90±29 vs 152±88 mg/dl in animals without and with tungsten treatment, respectively). In Figure 6, the efficient inhibition of xanthine oxidase by tungsten is shown in animals with and without feeding a high-fat and high-cholesterol diet over 35 days. Inhibition of xanthine oxidase in animals with hyperlipidemia was associated with a significant decrease in the number of ED1-positive cells infiltrating the cortical tubulointerstitium (Figure 7). Staining of renal tissues for ICAM-1 revealed no substantial changes between animals on a standard diet and hyperlipidemic rats with and without xanthine oxidase inhibition (not shown). Compared with the normolipemic controls, hyperlipidemic animals without xanthine oxidase inhibition showed intense and continuous circumferential staining for VCAM-1 at the endothelial side of interlobular arteries. Also, increased staining was present in the peritubular interstitium, most probably corresponding to the peritubular capillaries. In hyperlipidemic animals in which xanthine oxidase had been inhibited, increased peritubular VCAM-1 expression was clearly reduced and interlobular arteries showed discontinuous VCAM-1 staining at the endothelial side (Figure 8). MCP-1 staining in the peritubular interstitium was increased in hyperlipidemic animals as compared with the normolipemic controls. In hyperlipidemic animals with xanthine oxidase inhibition, MCP-1 staining was diminished, as shown in Figure 9.Figure 7Number of ED1-positive cells in the cortical tubulointerstitium of control animals with standard diet, either with (n=5) or without (n=8) administration of the xanthine oxidase inhibitor tungsten, and of hyperlipidemic animals (HLP) fed a fat- and cholesterol-enriched diet for 35 days, either with (n=11) or without (n=11) administration of tungsten.**P<0.01 vs control groups and hyperlipidemic animals with tungsten treatment.View Large Image Figure ViewerDownload (PPT)Figure 8(a and b) Immunohistology for VCAM-1 in the controls on a standard diet, (c and d) and in hyperlipidemic animals without (e and f) and with inhibition of xanthine oxidase. Controls show a very weak positivity for VCAM-1 (a) at the endothelial side of interlobular arteries and (b) in the peritubular area. In hyperlipidemic animals, (c) continuous circumferential staining of interlobular arteries and (d) intense staining of the peritubular area is present (arrows), besides increased glomerular staining. Hyperlipidemic animals with xanthine oxidase inhibition show (e) discontinuous VCAM-1 staining of interlobular arteries and (f; arrows) decreased VCAM-1 staining of the peritubular area. Original magnification: × 400.View Large Image Figure ViewerDownload (PPT)Figure 9Immunohistology for MCP-1 in the controls on a standard diet, and in hyperlipidemic animals without and with inhibition of xanthine oxidase. Compared with the controls, (a) which show weak staining for MCP-1 in the peritubular region, (b; arrows) hyperlipidemic animals without xanthine oxidase inhibition exhibit a pronounced MCP-1 staining in the peritubular interstitium. (c) This peritubular staining (arrow) is clearly decreased in hyperlipidemic animals with xanthine oxidase inhibition. Original magnification × 400.View Large Image Figure ViewerDownload (PPT) Based on recent observations in a model with long-standing hyperlipidemia in uninephrectomized rats with glomerulonephritis,9.Scheuer H. Gwinner W. Hohbach J. et al.Oxidant stress in hyperlipidemia-induced renal damage.Am J Physiol. 2000; 278: F63-F74PubMed Google Scholar this study examined whether xanthine oxidase has a role in the early phase of hyperlipidemia-mediated renal alterations. Different to previous experimental studies, which examined lipid-associated renal injury in accelerated models with uninephrectomy and glomerulonephritis,5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 7.Kasiske B.L. O'Donnell M.P. Cleary M.P. Keane W.F. Effects of reduced renal mass on tissue lipids and renal injury in hyperlipidemic rats.Kidney Int. 1989; 35: 40-47Abstract Full Text PDF PubMed Scopus (63) Google Scholar, 8.Lee H.S. Jeong J.Y. Kim B.C. et al.Dietary antioxidant inhibits lipoprotein oxidation and renal injury in experimental focal segmental glomerulosclerosis.Kidney Int. 1997; 51: 1151-1159Abstract Full Text PDF PubMed Scopus (83) Google Scholar, 9.Scheuer H. Gwinner W. Hohbach J. et al.Oxidant stress in hyperlipidemia-induced renal damage.Am J Physiol. 2000; 278: F63-F74PubMed Google Scholar we chose to study the isolated and early effects of diet-induced hyperlipidemia using rats without glomerular disease or reduced renal mass. Analysis of serum creatinine and protein excretion, and morphological evaluation of renal tissues confirmed that hyperlipidemic animals were, in fact, at an early stage of renal alterations. Obvious changes were limited to a moderate tubulointerstitial infiltration with ED1-positive cells, whereas tubular atrophy and tubulointerstitial fibrosis were absent. Similar subtle renal alterations have been reported previously in hyperlipidemic rats without coexisting glomerular disease or renal mass reduction.16.Joles J.A. Kunter U. Janssen U. et al.Early mechanisms of renal injury in hypercholesterolemic or hypertriglyceridemic rats.J Am Soc Nephrol. 2000; 11: 669-683PubMed Google Scholar Analysis of renal tissues in hyperlipidemic animals demonstrated an increased xanthine oxidase activity and generation of ROS. Apparently, induction of xanthine oxidase by hyperlipidemia particularly includes increased expression in proximal tubules as indicated by higher activities and higher protein and mRNA levels of the enzyme in this tissue fraction, besides the expression of xanthine oxidase in the area of peritubular capillaries observed in the immunohistological studies. The mechanisms by which hyperlipidemia induced xanthine oxidase in proximal tubules are unclear. A previous study in cultured human renal proximal tubular cells showed cellular uptake of non-oxidized lipoproteins with subsequent metabolism in a superoxide anion radical-dependent reaction. However, the involved oxidase was not specified.17.Ong A.C. Moorhead J.F. Tubular lipidosis: epiphenomenon or pathogenetic lesion in human renal disease?.Kidney Int. 1994; 45: 753-762Abstract Full Text PDF PubMed Scopus (37) Google Scholar Yet, studies in arterial vessels of rabbits with hypercholesterolemia demonstrated increased ROS generation by xanthine oxidase.18.Ohara Y. Peterson T.E. Harrison D.G. Hypercholesterolemia increases endothelial superoxide anion production.J Clin Invest. 1993; 91: 2546-2551Crossref PubMed Scopus (1608) Google Scholar, 19.Mugge A. Brandes R.P. Boger R.H. et al.Vascular release of superoxide radicals is enhanced in hypercholesterolemic rabbits.J Cardiovasc Pharmacol. 1994; 24: 994-998Crossref PubMed Scopus (126) Google Scholar Analysis of antioxidant enzymes revealed increased glutathione peroxidase activity in proximal tubular and tubulointerstitial samples. Such increased enzyme expression could represent a protective mechanism by which renal cells intercept the detrimental effects of lipid peroxides, as suggested by studies in cultured endothelial and renal proximal tubular cells.20.Thomas M.E. Schreiner G.F. Contribution of proteinuria to progressive renal injury: consequences of tubular uptake of fatty acid bearing albumin.Am J Nephrol. 1993; 13: 385-398Crossref PubMed Scopus (103) Google Scholar, 21.Thomas J.P. Geiger P.G. Girotti A.W. Lethal damage to endothelial cells by oxidized low density lipoprotein: role of selenoperoxidases in cytoprotection against lipid hydroperoxide- and iron-mediated reactions.J Lipid Res. 1993; 34: 479-490Abstract Full Text PDF PubMed Google Scholar Activities of SOD were unchanged, despite increased superoxide anion radical generation in renal samples of hyperlipidemic animals. Based on these results, it can be concluded that the antioxidant enzyme defense was not sufficiently upregulated by the increased load of ROS. Such inadequate response of the antioxidant enzymatic defense system to oxidative stress is in keeping with observations in several animal models of renal diseases as recently summarized.22.Gwinner W. Grone H.J. Role of reactive oxygen species in glomerulonephritis.Nephrol Dial Transplant. 2000; 15: 1127-1132Crossref PubMed Scopus (52) Google Scholar Hyperlipidemia has been reported to induce upregulation of the adhesion molecules ICAM-1 and VCAM-1 and of the chemokine MCP-1 in the kidney, factors that are important in the recruitment and attraction of macrophages.5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 11.Hattori M. Nikolic-Paterson D.J. Miyazaki K. et al.Mechanisms of glomerular macrophage infiltration in lipid-induced renal injury.Kidney Int. 1999; 56: S47-S50Abstract Full Text Full Text PDF Google Scholar, 12.Kodama N. Otani H. Yamada Y. et al.Involvement of MCP-1 and M-CSF in glomerular foam cell formation in ExHC rats.Kidney Int. 1999; 56: S174-S177Abstract Full Text PDF Google Scholar Moreover, the expression of these factors was shown to be regulated by ROS.13.Marui N. Offermann M.K. Swerlick R. et al.Vascular cell adhesion molecule-1 (VCAM-1) gene transcription and expression are regulated through an antioxidant-sensitive mechanism in human vascular endothelial cells.J Clin Invest. 1993; 92: 1866-1874Crossref PubMed Scopus (954) Google Scholar, 14.Lakshminarayanan V. Beno D.W. Costa R.H. Roebuck K.A. Differential regulation of interleukin-8 and intercellular adhesion molecule-1 by H2O2 and tumor necrosis factor-alpha in endothelial and epithelial cells.J Biol Chem. 1997; 272: 32910-32918Crossref PubMed Scopus (141) Google Scholar, 15.Satriano J.A. Shuldiner M. Hora K. et al.Oxygen radicals as second messengers for expression of the monocyte chemoattractant protein JE/MCP-1, and the monocyte colony-stimulating factor, CSF-1, in response to tumor necrosis factor-alpha and immunoglobulin G. Evidence for involvement of reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent oxidase.J Clin Invest. 1993; 92: 1564-1571Crossref PubMed Scopus (279) Google Scholar This led to the hypothesis that hyperlipidemia-mediated xanthine oxidase expression might induce these factors, which, in turn, facilitated the increased macrophage infiltration in the kidney. Therefore, experiments were performed with animals in which xanthine oxidase had been inhibited by administration of tungsten. Tungsten is an inhibitor that incorporates into the active center of the enzyme in place of molybdenum during enzyme synthesis, resulting in a non-functional xanthine oxidase.23.Johnson J.L. Waud W.R. Cohen H.J. Rajagopalan K.V. Molecular basis of the biological function of molybdenum. Molybdenum-free xanthine oxidase from livers of tungsten-treated rats.J Biol Chem. 1974; 249: 5056-5061Abstract Full Text PDF PubMed Google Scholar In hyperlipidemic animals with tungsten treatment, a profound inhibition of xanthine oxidase activity was documented in the cortical tubulointerstitium and in proximal tubules, along with a significant reduction of ED-1-positive cells in the cortical tubulointerstitium compared with hyperlipidemic animals without tungsten treatment. The reversal of this cell infiltration in hyperlipidemic animals with xanthine oxidase inhibition strongly suggests that xanthine oxidase indeed induces mediator pathways leading to the recruitment and attraction of infiltrating cells. Of the known mediators, expression of VCAM-1 and MCP-1 was found to be upregulated specifically in the peritubular area and for VCAM-1, in interlobular arteries, similar to previously reported results.5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 11.Hattori M. Nikolic-Paterson D.J. Miyazaki K. et al.Mechanisms of glomerular macrophage infiltration in lipid-induced renal injury.Kidney Int. 1999; 56: S47-S50Abstract Full Text Full Text PDF Google Scholar, 12.Kodama N. Otani H. Yamada Y. et al.Involvement of MCP-1 and M-CSF in glomerular foam cell formation in ExHC rats.Kidney Int. 1999; 56: S174-S177Abstract Full Text PDF Google Scholar Based on the reduced VCAM-1 and MCP-1 expression in hyperlipidemic animals, in which xanthine oxidase inhibition prevented the infiltration with ED1-positive cells, these two molecules may be of particular importance in this model. The essential role of MCP-1 in the recruitment of macrophages and development of atherosclerotic lesions has been convincingly demonstrated in hyperlipidemic mice genetically deficient in MCP-1.24.Gosling J. Slaymaker S. Gu L. et al.MCP-1 deficiency reduces susceptibility to atherosclerosis in mice that overexpress human apolipoprotein B.J Clin Invest. 1999; 103: 773-778Crossref PubMed Scopus (578) Google Scholar Once tubulointerstitial infiltration with macrophages is established, the further pathophysiological sequence may include local secretion of tumor growth factor-β and other mediators, eventually resulting in tubular atrophy and tubulointerstitial fibrosis as described earlier.5.Eddy A.A. Interstitial inflammation and fibrosis in rats with diet-induced hypercholesterolemia.Kidney Int. 1996; 50: 1139-1149Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 9.Scheuer H. Gwinner W. Hohbach J. et al.Oxidant stress in hyperlipidemia-induced renal damage.Am J Physiol. 2000; 278: F63-F74PubMed Google Scholar A pivotal question however is how these experimental observations apply to humans with hyperlipidemia. Cautious interpretation is necessary because of the known differences in lipid metabolism between rats and man. Rats have higher high-density lipoprotein cholesterol levels, and cholesterol- and fat-enriched diet induces increases in cholesterol-containing very low density lipoprotein and low-density lipoprotein particles.6.Grone H.J. Walli A. Grone E. et al.Induction of glomeru
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