Cloning Two Isoforms of Rat Cyclooxygenase: Differential Regulation of Their Expression

Li, Feng; Sun, Wenquin; Xia, Ying; Tang, Wei; Chanmugam, Prithiva; Soyoola, Emmanuel; Wilson, Cathal; Hwang, Dong Soo

doi:10.1006/abbi.1993.1601

Cited by 435 publications

(287 citation statements)

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“…COX-2 cDNA was synthesized by reverse-transcription PCR from rat microglia as discribed (Fiebich et al, 1996). The PCR product was sequenced and had no mutation relative to the published sequence of rat COX-2 (Feng et al, 1993). Total cellular RNA was extracted from confluent microglia and isolated by the acidic guanidium thiocyanate/phenol/chloroform method.…”

Section: Methodsmentioning

confidence: 99%

“…Two isoforms of cyclooxygenase, COX-1 and COX-2, have been identified, which, although they catalyze the same enzyme reaction, may have fundamentally different biological roles and distinct pharmacological profiles (Loll and Garavito, 1994;Feng et al, 1993). COX-I is constitutively expressed in most tissues and is thought to produce prostaglandins with more 'house-keeping' functions, such as gastric cytoprotection, vascular homeostasis and normal renal maintenance.…”

mentioning

confidence: 99%

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Expression and Regulation of Cyclooxygenase‐2 in Rat Microglia

Bauer

Lieb

Schulze‐Osthoff

et al. 1997

European Journal of Biochemistry

227

134

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Increased levels of prostanoids have been implicated in various neuropathological diseases, although little is known about their cellular sources inside the brain. In this study. we analyzed the expression of cyclooxygenase-2 (COX-2), a key enzyme in arachidonic acid metabolism, in rat microglia. COX-2 mRNA and protein as well as prostaglandin E, formation were almost undetectable in unstimulated microglial cultures but were found to be strongly upregulated in response to lipopolysaccharide. However, in contrast to most peripheral cells, proinflammatory cytokines such as tumor necrosis factor cr, interleukin-I [ j or interleukin-6 failed to markedly induce COX-2 expression. Similar effects were observed by analyzing transcription nuclear factor-lc B (NF-KB) which was strongly activated in microglia by lipopolysaccharide but not by incubation with cytokines. Moreover, known inhibitors of NF-KB activation, such as dexamethasone and the antioxidant pyrrolidine dithiocarbamate, as well as the protein kinase C (PKC) inhibitor Go6976, strongly reduced lipopolysaccharide-induced COX-2 transcription, indicating the involvement of NF-h-B and PKC in COX-2 expression. Our results suggest that microglia may represent an important source of prostanoids in the brain, thus reinforcing their prominent role in cerebral inflammatory processes.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Expression and Regulation of Cyclooxygenase‐2 in Rat Microglia

Bauer

Lieb

Schulze‐Osthoff

et al. 1997

European Journal of Biochemistry

227

134

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“…A 241-bp fragment of rat COX-2 cDNA produced by BamHI and EcoRI (from bp 409 to bp 650) was subcloned in pGEM4Z (20).…”

Section: Generation Of Rat Cox-1 and Cox-2 Riboprobe For Rnase Protecmentioning

confidence: 99%

A Role for Uric Acid in the Progression of Renal Disease

Kang

Nakagawa²,

Li³

et al. 2002

Journal of the American Society of Nephrology

1,157

770

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Abstract. Hyperuricemia is associated with renal disease, but it is usually considered a marker of renal dysfunction rather than a risk factor for progression. Recent studies have reported that mild hyperuricemia in normal rats induced by the uricase inhibitor, oxonic acid (OA), results in hypertension, intrarenal vascular disease, and renal injury. This led to the hypothesis that uric acid may contribute to progressive renal disease. To examine the effect of hyperuricemia on renal disease progression, rats were fed 2% OA for 6 wk after 5/6 remnant kidney (RK) surgery with or without the xanthine oxidase inhibitor, allopurinol, or the uricosuric agent, benziodarone. Renal function and histologic studies were performed at 6 wk. Given observations that uric acid induces vascular disease, the effect of uric acid on vascular smooth muscle cells in culture was also examined. RK rats developed transient hyperuricemia (2.7 mg/dl at week 2), but then levels returned to baseline by week 6 (1.4 mg/dl). In contrast, RKϩOA rats developed higher and more persistent hyperuricemia (6 wk, 3.2 mg/dl). Hyperuricemic rats demonstrated higher BP, greater proteinuria, and higher serum creatinine than RK rats. Hyperuricemic RK rats had more renal hypertrophy and greater glomerulosclerosis (24.2 Ϯ 2.5 versus 17.5 Ϯ 3.4%; P Ͻ 0.05) and interstitial fibrosis (1.89 Ϯ 0.45 versus 1.52 Ϯ 0.47; P Ͻ 0.05). Hyperuricemic rats developed vascular disease consisting of thickening of the preglomerular arteries with smooth muscle cell proliferation; these changes were significantly more severe than a historical RK group with similar BP. Allopurinol significantly reduced uric acid levels and blocked the renal functional and histologic changes. Benziodarone reduced uric acid levels less effectively and only partially improved BP and renal function, with minimal effect on the vascular changes. To better understand the mechanism for the vascular disease, the expression of COX-2 and renin were examined. Hyperuricemic rats showed increased renal renin and COX-2 expression, the latter especially in preglomerular arterial vessels. In in vitro studies, cultured vascular smooth muscle cells incubated with uric acid also generated COX-2 with time-dependent proliferation, which was prevented by either a COX-2 or TXA-2 receptor inhihbitor. Hyperuricemia accelerates renal progression in the RK model via a mechanism linked to high systemic BP and COX-2-mediated, thromboxane-induced vascular disease. These studies provide direct evidence that uric acid may be a true mediator of renal disease and progression.

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“…Hybridization was performed as previously described. 38 In brief, approximately 5 ϫ 10 5 recombinant phages from the cDNA library were screened with the 32 P-labeled oligonucleotide probe. Hybridization to filters was carried out overnight at 50°C in Hybrisol I (ONCOR, Norcross, GA), with 200 g/ml of heat-denatured, sheared salmon sperm DNA, 5 g/ml E. coli DNA, and a concentration of 2 ϫ 10 6 cpm/ml of the probe.…”

Section: Molecular Cloning Of Rat MDC and Ccr4mentioning

confidence: 99%