PubMed Health⌕ Search

Biomedical subjects

Raymond C Harris

Publications and source records attributed to Raymond C Harris.

48 records · Page 3Linked to original sources
PubMed2002

Cyclooxygenase-2 promotes early atherosclerotic lesion formation in LDL receptor-deficient mice.

BACKGROUND: Atherosclerosis has features of an inflammatory disease. Because cyclooxygenase (COX)-2 is expressed in atherosclerotic lesions and promotes inflammation, we tested the hypotheses that selective COX-2 inhibition would reduce early lesion formation in LDL receptor-deficient (LDLR-/-) mice and that macrophage COX-2 expression contributes to atherogenesis in LDLR-/- mice. METHODS AND RESULTS: Treatment of male LDLR-/- mice fed the Western diet with rofecoxib or indomethacin for 6 weeks resulted in significant reductions in atherosclerosis in the proximal aorta (25% and 37%) and in the aorta en face (58% and 57%), respectively. Rofecoxib treatment did not inhibit platelet thromboxane production, a COX-1-mediated process, but it significantly reduced the urinary prostacyclin metabolite 2,3-dinor-6-keto-PGF1alpha. Fetal liver cell transplantation was used to generate LDLR-/- mice null for expression of the COX-2 gene by macrophages. After 8 weeks on the Western diet, COX-2-/- --> LDLR-/- mice developed significantly less (33% to 39%) atherosclerosis than control COX-2+/+ --> LDLR-/- mice. In both the inhibitor studies and the transplant studies, serum lipids did not differ significantly between groups. CONCLUSIONS: The present studies provide strong pharmacological and genetic evidence that COX-2 promotes early atherosclerotic lesion formation in LDLR-/- mice in vivo. These results support the potential of anti-inflammatory approaches to the prevention of atherosclerosis.

Animals↗
PubMed2002

Cyclooxygenase-2 inhibition and renal physiology.

In the adult mammalian kidney, high levels of cyclooxygenase (COX)-2 expression can be detected in the macula densa and associated cortical thick ascending limb cells and medullary interstitial cells. In the renal cortex, COX-2 expression increases in high renin states, and selective COX-2 inhibitors significantly decrease plasma renin levels. In the medullary region of the kidney, the expression of COX-2 increases in response to a high-salt diet and water deprivation. The most important prostanoids in the kidney are prostaglandin (PG)I(2), or prostacyclin, and PGE(2). PGE(2) diminishes sodium reabsorption; thereby, its inhibition results in sodium retention that can manifest clinically in a variety of ways, such as peripheral edema, increased blood pressure (mainly in treated hypertensive patients), weight gain, and occasionally deterioration of heart failure. PGI(2) increases potassium secretion. As such, its inhibition can result in hyperkalemia, particularly in patients with underlying renal insufficiency. PGI(2) is also a potent vasodilator and helps maintain renal perfusion in conditions of decreased actual or effective circulating volume; its inhibition in such patients can result in acute renal failure. A variety of studies has been conducted to examine the effects of celecoxib and rofecoxib on renal function. These incorporate various study designs directly, making it virtually impossible to compare data across studies. It is apparent from such studies, coupled with published case reports, that the impact of both celecoxib and rofecoxib on renal function (including development of edema and hypertension) is similar to that of nonselective nonsteroidal anti-inflammatory drugs (NSAIDs). Studies comparing the 2 COX-2 inhibitors conflict in their interpretation. Overall, the data suggest similar effects on renal function among all NSAIDs when used at comparable doses.

Adolescent↗
PubMed2002

Role of membrane-bound heparin-binding epidermal growth factor-like growth factor (HB-EGF) in renal epithelial cell branching.

UNLABELLED: Role of membrane-bound heparin-binding epidermal growth factor-like growth factor (HB-EGF) in renal epithelial cell branching. BACKGROUND: The developing metanephros is characterized by growth and differentiation of the ureteric bud and the surrounding mesenchymal tissue. These processes can be influenced by several growth factors, including epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha). We examined whether another member of the EGF family of growth factors, heparin-binding epidermal growth factor (HB-EGF), might act as a morphogen in renal epithelial tubulogenesis. METHODS: Expression of HB-EGF mRNA and immunoreactive protein were examined in fetal, neonatal and adult rat kidneys. For in vitro studies of tubulogenesis, a rat renal epithelial cell line (NRK52E) stably transfected with proHB-EGF (NRKproHB-EGF) was treated with TPA for 30 minutes, washed with 2 mol/L NaCl to remove soluble HB-EGF trapped by cell surface heparan sulfate proteoglycan and replated onto plastic dishes in the absence of fetal calf serum. In further experiments, NRKproHB-EGF were suspended in a type I collagen gel in serum-free media. RESULTS: Northern blot analysis indicated that HB-EGF was strongly expressed in embryonic rat kidney (embryonic days 18-20) and was still increased in the neonatal kidney (day 10), compared to the low basal levels in adult kidney. Immunohistochemical analysis confirmed that immunoreactive HB-EGF expression in the fetal rat kidney was localized predominantly to the ureteric bud. When NRKproHB-EGF were plated onto plastic substrata, they became progressively flattened and enlarged and exhibited filopoidia. By 10 hours after plating, NRKproHB-EGF began to migrate and subsequently developed cell-cell contact and fully established tubular-like structures. Immunoelectron microscopy revealed that the initial recovery of cellular proHB-EGF was localized predominantly to areas of cell-cell attachment. No tubule-like structures were observed in similarly treated NRK52E cells transfected with the vector alone. In collagen gels, NRKproHB-EGF developed short tubule-like structures in the absence of TPA treatment, but with simultaneous TPA treatment, longer and more arborized structures developed. MMP-1 mRNA and immunoreactive protein increased in the TPA-treated cells, suggesting that protein kinase C-mediated collagenase activity was important for the observed tubulogenesis. However, inhibition of EGF receptor tyrosine kinase with AG 1478 significantly blunted the TPA-induced tubulogenesis by NRKproHB-EGF grown in collagen gels. CONCLUSIONS: These results indicate that membrane-bound HB-EGF can mediate both epithelial cell branching and cell motility. Localization of proHB-EGF to the site of cell-cell contact and development of tubule-like structures in collagen gels suggests that proHB-EGF may be an important morphogen for renal epithelial cells.

Animals↗
PubMed2002

Angiotensin II activates nuclear transcription factor-kappaB through AT1 and AT2 receptors.

BACKGROUND: Recent evidence suggests that angiotensin II (Ang II) induces a variety of proinflammatory mediators including chemokines. Nuclear factor-kappaB (NF-kappaB) activation plays an important role in Ang II-mediated inflammation. The present study investigated which Ang II receptor subtype is involved in NF-kappaB activation. We focused particularly on the Ang II subtype 2 (AT2) receptor because we previously observed that Ang II-induction of the chemokine RANTES in vitro and in vivo is mediated through AT2 receptors. METHODS: AT1 or AT2 receptors were selectively overexpressed in COS7 cells that normally do not express Ang II receptors. In addition, rat glomerular endothelial cells (GER) that express AT1 and AT2 receptors and PC12 cells that exclusively exhibit AT2 receptors were studied also. Ang II-receptor expression was confirmed by Western blots of membrane lysates. NF-kappaB DNA binding in vitro was detected by electrophoretic shift assays. In addition, in vivo transactivation of a reporter gene construct with kappa enhancer coupled to luciferase also was investigated. Expression of the inhibitor of kappaB alpha (IkappaB-alpha) was detected by Western blots. RESULTS: In AT1 or AT2 receptor transfected cells, but not untransfected COS7 cells, 10-7 mol/L Ang II induced NF-kappaB DNA binding in vitro, as detected by electrophoretic shift assays and in vivo transactivation of a reporter gene construct. The AT2 receptor antagonist PD 123319 but not losartan attenuated Ang II-mediated NF-kappaB activation in COS7 cells transfected with AT2 receptors. While Ang II also induced NF-kappaB activation in PC12 cells, this activation was blocked by PD 123319. Finally, stimulation of GERs with Ang II led to the activation of NF-kappaB through both subtypes of Ang II receptors. Nuclear extracts from COS7 cells transfected with AT2 receptors and PC12 cells with NF-kappaB DNA-binding activity consisted of p50/p65 complexes. There was no difference in subunit composition of nuclear proteins from Ang II-stimulated AT1 receptor transfected COS7 cells. An artificial peptide (p-Amino-Phe6-Ang II) with a high affinity for the AT2 receptor also activated NF-kappaB. Ang II-induced activation of NF-kappaB was associated with degradation of IkappaB-alpha in all studied cell lines. CONCLUSIONS: Our results clearly demonstrate in various cell lines that Ang II induces NF-kappaB activation through AT2 receptors. These data may have important therapeutic consequences, because potential Ang II-mediated proinflammatory renal and cardiovascular effects may not be totally antagonized by the currently increased clinical use of AT1 receptor antagonists.

Angiotensin II↗
PubMed2002

Cyclooxygenase-2 inhibitor blocks expression of mediators of renal injury in a model of diabetes and hypertension.

BACKGROUND: We previously reported that renal cortical cyclooxygenase (COX-2) expression increased following subtotal nephrectomy, and chronic treatment with a selective COX-2 inhibitor, SC58236, reduced proteinuria and retarded the development of glomerulosclerosis. The present studies were designed to examine the effects of COX-2 inhibition in a model of diabetic nephropathy. METHODS: Rats were divided into three groups: control, diabetic (streptozotocin-induced diabetic animals with superimposed DOCA/salt hypertension; right nephrectomy and DOCA treatment), and treated (administration of the selective COX-2 inhibitor, SC58236, to a subset of diabetic/DOCA/salt rats). Insulin was administered to maintain blood glucose in the 200 to 300 mg/dL range. RESULTS: Systolic blood pressure in the two diabetic groups was elevated within one week and remained elevated until sacrifice at six weeks (control, 108 +/- 2 mm Hg; diabetic, 158 +/- 4 mm Hg; treated, 156 +/- 7 mm Hg). When measured at six weeks, immunoreactive COX-2 expression in the renal cortex of the diabetic rats was 2.5 +/- 0.3-fold of control animals (N = 7). Immunohistochemical localization indicated increased expression in macula densa and surrounding cortical thick ascending limb of Henle (cTALH). The COX-2 inhibitor decreased COX-2 expression in diabetic rats to 1.3 +/- 0.1-fold control. In addition, SC58236 decreased expression of PAI-1 (diabetic vs. treated, 3.2 +/- 0.5 vs. 1.7 +/- 0.2-fold control, N = 7, P < 0.05), vascular endothelial growth factor (VEGF; 2.0 +/- 0.2 vs. 1.2 +/- 0.2; N = 7, P < 0.05), fibronectin (2.4 +/- 0.3 to 1.3 +/- 0.1; N = 7, P < 0.05) and transforming growth factor-beta (TGF-beta; 2.1 +/- 0.2 vs. 1.3 +/- 0.2; N = 7, P < 0.05). Proteinuria at six weeks was decreased in the SC58236-treated rats (149 +/- 8 vs. 92 +/- 8 mg/24 h; N = 7, P < 0.01). The mesangial sclerosis index, defined as increases in extracellular matrix within the mesangial space, was determined at six weeks; the control group had an index of 0.06 +/- 0.01, the diabetic group was 2.7 +/- 0.04 and the treated group was 0.6 +/- 0.03 (P < 0.0001 compared to the diabetic group). CONCLUSIONS: These results suggest that in an experimental model of diabetes and hypertension, inhibition of COX-2 expression decreases potential mediators of glomerular and tubulointerstitial injury and also decreases biochemical, functional and structural markers of renal injury.

Animals↗
PubMed2002

Prostaglandins that increase renin production in response to ACE inhibition are not derived from cyclooxygenase-1.

It is well known that nonselective, nonsteroidal anti-inflammatory drugs inhibit renal renin production. Our previous studies indicated that angiotensin-converting enzyme inhibitor (ACEI)-mediated renin increases were absent in rats treated with a cyclooxygenase (COX)-2-selective inhibitor and in COX-2 -/- mice. The current study examined further whether COX-1 is also involved in mediating ACEI-induced renin production. Because renin increases are mediated by cAMP, we also examined whether increased renin is mediated by the prostaglandin E(2) receptor EP(2) subtype, which is coupled to G(s) and increases cAMP. Therefore, we investigated if genetic deletion of COX-1 or EP(2) prevents increased ACEI-induced renin expression. Age- and gender-matched wild-type (+/+) and homozygous null mice (-/-) were administered captopril for 7 days, and plasma and renal renin levels and renal renin mRNA expression were measured. There were no significant differences in the basal level of renal renin activity from plasma or renal tissue in COX-1 +/+ and -/- mice. Captopril administration increased renin equally [plasma renin activity (PRA): +/+ 9.3 +/- 2.2 vs. 50.1 +/- 10.9; -/- 13.7 +/- 1.5 vs. 43.9 +/- 6.6 ng ANG I x ml(-1) x h(-1); renal renin concentration: +/+ 11.8 +/- 1.7 vs. 35.3 +/- 3.9; -/- 13.0 +/- 3.0 vs. 27.8 +/- 2.7 ng ANG I x mg protein(-1) x h(-1); n = 6; P < 0.05 with or without captopril]. ACEI also increased renin mRNA expression (+/+ 2.4 +/- 0.2; -/- 2.1 +/- 0.2 fold control; n = 6-10; P < 0.05). Captopril led to similar increases in EP(2) -/- compared with +/+. The COX-2 inhibitor SC-58236 blocked ACEI-induced elevation in renal renin concentration in EP(2) null mice (+/+ 24.7 +/- 1.7 vs. 9.8 +/- 0.4; -/- 21.1 +/- 3.2 vs. 9.3 +/- 0.4 ng ANG I x mg protein(-1) x h(-1); n = 5) as well as in COX-1 -/- mice (SC-58236-treated PRA: +/+ 7.3 +/- 0.6; -/- 8.0 +/- 0.9 ng ANG I x ml(-1) x h(-1); renal renin: +/+ 9.1 +/- 0.9; -/- 9.6 +/- 0.5 ng ANG I x mg protein(-1) x h(-1); n = 6-7; P < 0.05 compared with no treatment). Immunohistochemical analysis of renin expression confirmed the above results. This study provides definitive evidence that metabolites of COX-2 rather than COX-1 mediate ACEI-induced renin increases. The persistent response in EP(2) nulls suggests involvement of prostaglandin E(2) receptor subtype 4 and/or prostacyclin receptor (IP).

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2002

Mineralocorticoid regulation of cyclooxygenase-2 expression in rat renal medulla.

The renal inner medulla and its distal one-third, the papilla, are major sites of prostanoid synthesis involved in water and electrolyte homeostasis. These sites contain variable levels of cyclooxygenase (COX)-2, a key prostaglandin synthase enzyme that is sensitive to adrenal steroids. Immunoreactive renal medullary COX-2, restricted to interstitial cells in control adult rats, shows a gradient of intense staining at the tip of the papilla that gradually diminishes to undetectable levels in the proximal inner medulla. We used adrenalectomy (ADX) and steroid replacement to investigate the effects of steroids on papillary COX-2. Immunoblots demonstrate that papillary COX-2 was reduced by one-half after 2 wk ADX; glucocorticoid replacement ameliorated the decline but not to control levels. Mineralocorticoid (deoxycorticosterone acetate; DOCA) replacement stimulated papillary COX-2 more than fivefold over control; both the intensity of immunostaining and the numbers of COX-2-positive cells in the inner medulla increased. Similar stimulation of papillary COX-2 resulted from DOCA treatment of normal control rats, but the response was blunted in rats fed a low-salt diet and absent in Brattleboro rats. DOCA treatment of mouse renal medullary interstitial cells in culture had no effect, but increased tonicity of the culture medium with NaCl caused strong upregulation of COX-2. Urea, a permeant molecule, had no effect. Together, these results suggest that mineralocorticoids lead to upregulation of COX-2 in rat renal medulla by indirect pathways, probably involving induced electrolyte hypertonicity in the interstitial fluid.

Adrenalectomy↗
PubMed2002

Cyclooxygenase-2 and the kidney: functional and pathophysiological implications.

In adult mammalian kidney, cyclooxygenase-2 (COX-2) expression is found in restricted subpopulations of cells. High levels of expression can be detected in the macula densa (MD) and associated cortical thick ascending limb of Henle (cTALH) cells and medullary interstitial cells (MICs). In human biopsy specimens, COX-2 expression is also detected in glomerular podocytes and increased podocyte expression is seen in experimental models of progressive glomerular injury. Physiological regulation of COX-2 in these cellular compartments suggests functional roles for eicosanoid products of the enzyme. COX-2 expression increases in high-renin states (salt restriction, angiotensin-converting enzyme inhibition, renovascular hypertension) and selective COX-2 inhibitors significantly decrease plasma renin levels, renal renin activity and mRNA expression. There is evidence for negative regulation of MD/cTALH COX-2 by angiotensin II and by glucocorticoids and mineralocorticoids. Conversely, nitric oxide (NO) generated by neuronal nitric oxide synthase (nNOS) is a positive modulator of COX-2 expression. Decreased extracellular chloride increases COX-2 expression in cultured cTALH, an effect mediated by increased p38 MAP kinase activity and, in vivo, a sodium-deficient diet increases expression of activated p38 in MD/cTALH. In contrast to COX-2 in MD/cTALH, COX-2 expression in MICs increases in response to a high-salt diet, as well as water deprivation. Studies in cultured MICs confirm that expression is increased in response to hypertonicity, and expression is mediated at least in part by nuclear factor-kappaB (NFkappaB) activation. COX-2 inhibition leads to apoptosis of MICs in response to hypertonicity in vitro and following water deprivation in vivo. In addition, COX-2 metabolites appear to be important mediators of medullary blood flow and renal salt handling. Therefore, there is increasing evidence that COX-2 is an important physiological mediator of kidney function.

Animals↗
PubMed2001

Induction of collecting duct morphogenesis in vitro by heparin-binding epidermal growth factor-like growth factor.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF), a member of the epidermal growth factor family of growth factors, is synthesized as a membrane-an-chored precursor (proHB-EGF) that is capable of stimulating adjacent cells in a juxtacrine manner. ProHB-EGF is cleaved in a protein kinase C-dependent process, to yield the soluble form. It was observed that HB-EGF acts as a morphogen for the collecting duct system in developing kidneys. HB-EGF protein was expressed in the ureteric bud of embryonic kidneys. Cultured mouse ureteric bud cells (UBC) produced HB-EGF via protein kinase C activation. After stimulation with phorbol ester (12-O-tetradecanoylphorbol-13-acetate) or recombinant soluble HB-EGF, UBC cultured in three-dimensional collagen gels formed short tubules with varied abundant branches. When proHB-EGF-transfected UBC were stimulated with 12-O-tetradecanoylphorbol-13-acetate and cultured in collagen gels, they exhibited linear growth, forming long tubular structures with few branches at the time of appearance of proHB-EGF on the cell surface. The structures exhibited a strong resemblance to the early branching ureteric bud of embryonic kidneys. When UBC were cultured in the presence of transforming growth factor-beta and soluble HB-EGF, they formed long tubules and few branches, similar to the structures observed in proHB-EGF-transfected UBC. These cells exhibited apical-basolateral polarization and expression of the water channel aquaporin-2. These findings indicate that soluble HB-EGF and proHB-EGF induce branching tubulogenesis in UBC in different ways. Juxtacrine activation by proHB-EGF or the synergic action of soluble HB-EGF with transforming growth factor-beta is important for well balanced morphogenesis of the collecting duct system.

Animals↗
PubMed2000

Ischemic acute renal failure induces differential expression of small heat shock proteins.

AlphaB-crystallin and heat shock protein (hsp) 25 are structurally and functionally related small stress proteins induced by a variety of insults, including heat and ischemia. Cytoprotection by these two hsp is thought to result from molecular chaperoning and/or cytoskeletal stabilization. Because renal ischemia is characterized by disruption of the renal tubular cell actin cytoskeleton, this study was conducted to determine the localization and quantify the expression and phosphorylation of both hsp in renal cortex, isolated glomeruli, outer medulla, and inner medulla of rats after bilateral renal ischemia. Sham-operated kidneys had similarly small amounts of hsp25 and alphaB-crystallin in cortex and glomeruli, with substantially greater amounts of alphaB-crystallin versus hsp25 in outer and inner medulla. Ischemia resulted in significantly increased hsp25 (and hsp70i) but variable alphaB-crystallin levels in cortex and outer medulla, and progressively decreased glomerular hsp25 phosphorylation. In sham-operated kidneys, hsp25 localized to glomeruli, vessels, and collecting ducts, with alphaB-crystallin primarily in medullary thin limbs and collecting ducts. After ischemia, hsp25 accumulated in proximal tubules in cortex and outer medulla, while alphaB-crystallin labeling became nonhomogeneous in outer medulla, and increased in Bowman's capsule. It is concluded that: (1) There is striking differential expression of hsp25 and alphaB-crystallin in various renal compartments; and (2) Renal ischemia results in differential accumulation of hsp25 and alphaB-crystallin, with hsp25 part of a generalized stress response in renal proximal tubular cells, which may play a role in recovery from ischemia-induced actin filament disruption.

Acute Kidney Injury↗
PubMedDate not supplied

EP24.15 interacts with the angiotensin II type I receptor and bradykinin B2 receptor.

The carboxyl-terminal cytoplasmic domain of the angiotensin II type 1 receptor (AT1) is known to interact with several classes of intracellular proteins that may modulate receptor function. Employing yeast two-hybrid screening of a human embryonic kidney cDNA library with the carboxyl-terminal cytoplasmic domain of the AT1 receptor as a bait, we have isolated EP24.15 (EC 3.4.24.15, thimet oligopeptidase) as a potentially interacting protein. EP24.15 is widely distributed and is known to degrade bioactive peptides such as angiotensin I and II and bradykinin. In addition, EP24.15 was previously identified as a putative soluble angiotensin II binding protein. Two-hybrid screening also determined that EP24.15 can interact with the B2 bradykinin receptor. Transient expression of EP24.15 in a porcine kidney epithelial cell line stably expressing full length AT1 and full length B2 followed by affinity chromatography and co-immunoprecipitation confirmed EP24.15 association with both AT1 and B2 receptors. EP24.15 was also co-immunoprecipitated with AT1 and B2 in rat kidney brush border membranes (BBM) and basolateral membranes (BLM). Both AT1 and B2 undergo ligand-induced endocytosis. Analysis of endosomal fractions following immunoprecipitation with AT1 or B2 antibodies detected strong association of EP24.15 with the receptors in both light and heavy endosomal populations. Therefore, the present study indicates that EP24.15 associates with AT1 and B2 receptors both at the plasma membrane and after receptor internalization and suggests a possible mechanism for endosomal disposition of ligand that may facilitate receptor recycling.

Animals↗