Coinduction of Nitric-oxide Synthase and Arginase I in Cultured Rat Peritoneal Macrophages and Rat Tissues in Vivo by Lipopolysaccharide

Nitric oxide is synthesized by nitric-oxide synthase from arginine, a common substrate of arginase. Rat peritoneal macrophages were cultured in the presence of bacterial lipopolysaccharide (LPS), and expression of the inducible isoform of nitric-oxide synthase (iNOS) and liver-type arginase (arginase I) was analyzed. mRNAs for iNOS and arginase I were induced by LPS in a dose-dependent manner. iNOS mRNA appeared 2 h after LPS treatment and increased to a near maximum at 8-12 h. On the other hand, arginase I mRNA that was undetectable prior to the treatment began to increase after 4 h with a lag time and reached a maximum at 12 h. Immunoblot analysis showed that iNOS and arginase I proteins were also induced. mRNA for arginase II, an arginase isozyme, was not detected in the LPS-activated peritoneal cells. mRNA for CCAAT/enhancer-binding protein β (C/EBPβ), a transactivator of the arginase I gene, was also induced, and the induction was more rapid than that of arginase I mRNA. Changes in iNOS and arginase I mRNAs were also examined in LPS-injected rats in vivo iNOS mRNA increased rapidly in the lung and spleen, reached a maximum 2-6 h after the LPS treatment, and decreased thereafter. Arginase I mRNA was induced markedly and more slowly in both tissues, reaching a maximum in 12 h. Thus, arginase I appears to have an important role in down-regulating nitric oxide synthesis in murine macrophages by decreasing the availability of arginine, and the induction of arginase I is mediated by C/EBPβ. Nitric oxide is synthesized by nitric-oxide synthase from arginine, a common substrate of arginase. Rat peritoneal macrophages were cultured in the presence of bacterial lipopolysaccharide (LPS), and expression of the inducible isoform of nitric-oxide synthase (iNOS) and liver-type arginase (arginase I) was analyzed. mRNAs for iNOS and arginase I were induced by LPS in a dose-dependent manner. iNOS mRNA appeared 2 h after LPS treatment and increased to a near maximum at 8-12 h. On the other hand, arginase I mRNA that was undetectable prior to the treatment began to increase after 4 h with a lag time and reached a maximum at 12 h. Immunoblot analysis showed that iNOS and arginase I proteins were also induced. mRNA for arginase II, an arginase isozyme, was not detected in the LPS-activated peritoneal cells. mRNA for CCAAT/enhancer-binding protein β (C/EBPβ), a transactivator of the arginase I gene, was also induced, and the induction was more rapid than that of arginase I mRNA. Changes in iNOS and arginase I mRNAs were also examined in LPS-injected rats in vivo iNOS mRNA increased rapidly in the lung and spleen, reached a maximum 2-6 h after the LPS treatment, and decreased thereafter. Arginase I mRNA was induced markedly and more slowly in both tissues, reaching a maximum in 12 h. Thus, arginase I appears to have an important role in down-regulating nitric oxide synthesis in murine macrophages by decreasing the availability of arginine, and the induction of arginase I is mediated by C/EBPβ. INTRODUCTIONNitric oxide (NO) 1The abbreviations used are: NOnitric oxideNOSnitric-oxide synthaseiNOSinducible isoform of NOSLPSlipopolysaccharidekbkilobase(s)C/EBPβCCAAT/enhancer-binding protein β. is a major molecule regulating blood vessel dilatation and immune response and functions as a neurotransmitter in the brain and peripheral nervous system (see Refs. 1Nathan C. Xie Q. J. Biol. Chem. 1994; 269: 13725-13728Abstract Full Text PDF PubMed Google Scholar, 2Schmidt H.H.H.W. Walter U. Cell. 1994; 78: 919-925Abstract Full Text PDF PubMed Scopus (1495) Google Scholar, 3Knowles R.G. Moncada S. Biochem. J. 1994; 298: 249-258Crossref PubMed Scopus (2485) Google Scholar for reviews). NO is synthesized from arginine by nitric-oxide synthase (NOS), generating citrulline. Cellular NO production is absolutely dependent on the availability of arginine. This amino acid can be obtained from exogenous sources via the blood circulation, from intracellular protein degradation, or by the endogenous synthesis of arginine. Major sites of arginine synthesis in ureotelic animals are the liver, where arginine generated in the urea cycle (ornithine cycle) is rapidly converted to urea and ornithine by arginase, and the kidney, where arginine is synthesized from citrulline and released into the blood circulation (see Ref. 4Morris S.M. Annu. Rev. Nutr. 1994; 12: 81-101Crossref Scopus (214) Google Scholar for a review). In other tissues and cell types, arginine can be generated from citrulline, which is produced as a coproduct of the NOS reaction, forming a cycle that is composed of NOS, argininosuccinate synthetase, and argininosuccinate lyase and that is termed the “citrulline-NO cycle” (5Hecker M. Sessa W.C. Harris H.J. Ånggárd E.E. Vane J.R. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 8612-8616Crossref PubMed Scopus (403) Google Scholar, 6Mitchell J.A. Hecker M. Vane J.R. Eur. J. Pharmacol. 1990; 176: 253-254Crossref PubMed Scopus (48) Google Scholar, 7Wu G. Brosnan J.T. Biochem. J. 1992; 281: 45-48Crossref PubMed Scopus (161) Google Scholar, 8Nussler A.K. Billiar T.R. Liu Z.-Z. Morris Jr., S.M. J. Biol. Chem. 1994; 269: 1257-1261Abstract Full Text PDF PubMed Google Scholar, 9Hattori Y. Campbell E.B. Gross S.S. J. Biol. Chem. 1994; 269: 9405-9408Abstract Full Text PDF PubMed Google Scholar, 10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). The inducible isoform of NOS (iNOS) and argininosuccinate synthetase are coinduced in activated murine macrophage-like RAW 264.7 cells (8Nussler A.K. Billiar T.R. Liu Z.-Z. Morris Jr., S.M. J. Biol. Chem. 1994; 269: 1257-1261Abstract Full Text PDF PubMed Google Scholar), in cultured vascular smooth muscle cells (9Hattori Y. Campbell E.B. Gross S.S. J. Biol. Chem. 1994; 269: 9405-9408Abstract Full Text PDF PubMed Google Scholar), and in vivo (10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar, 11Hattori Y. Shimoda S. Gross S.S. Biochem. Biophys. Res. Commun. 1995; 215: 148-153Crossref PubMed Scopus (28) Google Scholar). Argininosuccinate lyase is also induced in vivo (10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar, 11Hattori Y. Shimoda S. Gross S.S. Biochem. Biophys. Res. Commun. 1995; 215: 148-153Crossref PubMed Scopus (28) Google Scholar).On the other hand, arginine is utilized for both the arginase and NOS reactions. Thus, these two enzymes compete for arginine. At least two isoforms of arginase are present. Liver-type arginase (arginase I) is expressed almost exclusively in the liver and catalyzes the final step of urea synthesis. Arginase I activity (12Lamers W.H. Mooren P.G. De Graaf A. Charles R. Eur. J. Biochem. 1985; 146: 475-480Crossref PubMed Scopus (42) Google Scholar) and mRNA (13Morris Jr., S.M. Kepka D.M. Sweeney Jr., W.E. Avner E.D. Arch. Biochem. Biophys. 1989; 269: 175-180Crossref PubMed Scopus (34) Google Scholar) in rat liver increase markedly in the perinatal period, in coordination with other urea cycle enzymes. The enzyme is regulated by dietary protein (14Schimke R.T. J. Biol. Chem. 1962; 237: 459-468Abstract Full Text PDF PubMed Google Scholar) and hormones (15Nebes V.L. Morris Jr., S.M. Mol. Endocrinol. 1988; 2: 444-451Crossref PubMed Scopus (89) Google Scholar). Arginase I consists of three identical subunits of about 35,000 Da. cDNA clones were isolated from rat (16Dizikes G.J. Spector E.B. Cederbaum S.D. Somatic Cell Mol. Genet. 1986; 12: 375-384Crossref PubMed Scopus (43) Google Scholar, 17Kawamoto S. Amaya Y. Murakami K. Tokunaga F. Iwanaga S. Kobayashi K. Saheki T. Kimura S. Mori M. J. Biol. Chem. 1987; 262: 6280-6283Abstract Full Text PDF PubMed Google Scholar) and human (18Dizikes G.J. Grody W.W. Kern R.M. Cederbaum S.D. Biochem. Biophys. Res. Commun. 1986; 141: 53-59Crossref PubMed Scopus (68) Google Scholar, 19Haraguchi Y. Takiguchi M. Amaya Y. Kawamoto S. Matsuda I. Mori M. Proc. Natl. Acad. Sci. U. S. A. 1987; 84: 412-415Crossref PubMed Scopus (114) Google Scholar) liver. The rat (20Ohtake A. Takiguchi M. Shigeto Y. Amaya Y. Kawamoto S. Mori M. J. Biol. Chem. 1988; 263: 2245-2249Abstract Full Text PDF PubMed Google Scholar) and human (21Takiguchi M. Haraguchi Y. Mori M. Nucleic Acids Res. 1988; 16: 8789-8802Crossref PubMed Scopus (49) Google Scholar) genes are 11.5-12 kb long and consist of 8 exons. Promoter and enhancer regions of the rat gene were characterized (22Takiguchi M. Mori M. J. Biol. Chem. 1991; 266: 9186-9193Abstract Full Text PDF PubMed Google Scholar, 23Gotoh T. Haraguchi Y. Takiguchi M. Mori M. J. Biochem. 1994; 115: 778-788Crossref PubMed Scopus (26) Google Scholar, 24Chowdhury S. Gotoh T. Mori M. Takiguchi M. Eur. J. Biochem. 1996; 236: 500-509Crossref PubMed Scopus (32) Google Scholar). In addition to arginase I, an isozyme (arginase II) is present in extrahepatic tissues, including kidney, small intestine, and lactating mammary gland (25Glass R.D. Knox W.E. J. Biol. Chem. 1973; 248: 5785-5789Abstract Full Text PDF PubMed Google Scholar, 26Kaysen G.A. Strecker H.J. Biochem. J. 1973; 133: 779-788Crossref PubMed Scopus (127) Google Scholar, 27Herzfeld A. Raper S.M. Biochem. J. 1976; 153: 469-478Crossref PubMed Scopus (144) Google Scholar). The coinduction of NOS and arginase II activities in RAW 264.7 cells activated by lipopolysaccharide (LPS) was reported (28Wang W.W. Jenkinson C.P. Griscavage J.M. Kern R.M. Arabolos N.S. Byrns R.E. Cederbaum S.D. Ignarro L.J. Biochem. Biophys. Res. Commun. 1995; 210: 1009-1016Crossref PubMed Scopus (204) Google Scholar). We isolated cDNA for arginase II and showed that it is 59% identical with arginase I on the amino acid level (29Gotoh T. Sonoki T. Nagasaki A. Terada K. Takiguchi M. Mori M. FEBS Lett. 1996; 395: 119-122Crossref PubMed Scopus (210) Google Scholar). We also found that the enzyme is mitochondrial and that its mRNA is induced by LPS, dexamethasone, and cyclic adenosine monophosphate and is reduced by interferon-γ (29Gotoh T. Sonoki T. Nagasaki A. Terada K. Takiguchi M. Mori M. FEBS Lett. 1996; 395: 119-122Crossref PubMed Scopus (210) Google Scholar). To better understand the role of arginase isoforms in NO synthesis, we examined expression of the isoforms and iNOS in cultured rat peritoneal cells and in LPS-treated rats using RNA blots, immunoblots, and immunocytochemical analyses. We report here that iNOS and arginase I mRNAs and proteins are coinduced by LPS in cultured rat peritoneal macrophages and in the lung and spleen in vivo The induction of mRNA for CCAAT/enhancer-binding protein β (C/EBPβ), a transactivator of the arginase I gene, is also described.DISCUSSIONNO synthesis is regulated by depending on the availability of arginine, substrate of the NOS reaction, as well as by NOS activity and other factors. Arginine can be obtained via the blood circulation or by endogenous synthesis from citrulline. Arginine transport into cultured macrophages increases in response to LPS and to interferon-γ (35Bogle R.G. Baydoun A.R. Pearson J.D. Moncada S. Mann G.E. Biochem. J. 1992; 284: 15-18Crossref PubMed Scopus (212) Google Scholar, 36Sato H. Fujiwara M. Bannai S. J. Leukocyte Biol. 1992; 52: 161-164Crossref PubMed Scopus (49) Google Scholar). The cat-2 gene of the arginine transporter in cultured vascular smooth muscle cells is stimulated by interleukin-1β and by tumor necrosis factor-2 (37Gill D.J. Low B.C. Grigor M.R. J. Biol. Chem. 1996; 271: 11280-11283Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar). Furthermore, argininosuccinate synthetase and argininosuccinate lyase, which together synthesize arginine from citrulline, are induced in stimulated murine macrophages (8Nussler A.K. Billiar T.R. Liu Z.-Z. Morris Jr., S.M. J. Biol. Chem. 1994; 269: 1257-1261Abstract Full Text PDF PubMed Google Scholar, 10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar) and aortic smooth muscle cells (9Hattori Y. Campbell E.B. Gross S.S. J. Biol. Chem. 1994; 269: 9405-9408Abstract Full Text PDF PubMed Google Scholar), and intracellular arginine synthesis is enhanced.Activity of arginase that degrades arginine is also induced in activated macrophages (38Kung J.T. Brooks S.B. Jakway J.P. Leonard L.L. Talmage D.W. J. Exp. Med. 1977; 146: 665-672Crossref PubMed Scopus (141) Google Scholar, 39Currie G.A. Nature. 1978; 273: 758-759Crossref PubMed Scopus (246) Google Scholar). Arginase induced in activated RAW 264.7 cells, a mouse macrophage-like cell line, was shown to be arginase II (28Wang W.W. Jenkinson C.P. Griscavage J.M. Kern R.M. Arabolos N.S. Byrns R.E. Cederbaum S.D. Ignarro L.J. Biochem. Biophys. Res. Commun. 1995; 210: 1009-1016Crossref PubMed Scopus (204) Google Scholar, 29Gotoh T. Sonoki T. Nagasaki A. Terada K. Takiguchi M. Mori M. FEBS Lett. 1996; 395: 119-122Crossref PubMed Scopus (210) Google Scholar). On the other hand, in the present work, we noted that arginase I, not arginase II, is induced by LPS in primary cultured rat peritoneal macrophages and in the lung and spleen in vivo This was unexpected because arginase I has been thought to be expressed almost exclusively in the liver. Differential induction of the two arginase isoforms in the mouse macrophage-like cell line and in primary cultured rat macrophages may be due to differences in animal species or to differences between the established cell line and the primary cultured cells. Arginase I mRNA was induced more slowly than iNOS mRNA. The induced arginase I probably decreases arginine availability for the NOS reaction in activated macrophages and may down-regulate the overproduction of NO. All of these results suggest that there is a complex regulation of genes encoding enzymes and transporter proteins involved in arginine metabolism that together control NO production in cells.Arginase I mRNA was induced slowly in cultured peritoneal macrophages and also in vivo with an apparent lag time of a few hours. Therefore, the induction of arginase I appears to be mediated by a transcription factor(s) that is synthesized de novo in response to LPS stimulation. C/EBP family members bind to the promoter and enhancer regions of the arginase I gene and activate the promoter (Refs. 22Takiguchi M. Mori M. J. Biol. Chem. 1991; 266: 9186-9193Abstract Full Text PDF PubMed Google Scholar, 23Gotoh T. Haraguchi Y. Takiguchi M. Mori M. J. Biochem. 1994; 115: 778-788Crossref PubMed Scopus (26) Google Scholar, 24Chowdhury S. Gotoh T. Mori M. Takiguchi M. Eur. J. Biochem. 1996; 236: 500-509Crossref PubMed Scopus (32) Google Scholar; see Ref. 40Takiguchi M. Mori M. Biochem. J. 1995; 312: 649-659Crossref PubMed Scopus (97) Google Scholar for a review), C/EBPβ being the most potent. The present study also shows that C/EBPβ mRNA is induced in the LPS-stimulated peritoneal macrophages. The induction of C/EBPβ mRNA is more rapid than that of arginase I mRNA. Thus, the induction of arginase I appears to be mediated, at least in part, by the induction of C/EBPβ. INTRODUCTIONNitric oxide (NO) 1The abbreviations used are: NOnitric oxideNOSnitric-oxide synthaseiNOSinducible isoform of NOSLPSlipopolysaccharidekbkilobase(s)C/EBPβCCAAT/enhancer-binding protein β. is a major molecule regulating blood vessel dilatation and immune response and functions as a neurotransmitter in the brain and peripheral nervous system (see Refs. 1Nathan C. Xie Q. J. Biol. Chem. 1994; 269: 13725-13728Abstract Full Text PDF PubMed Google Scholar, 2Schmidt H.H.H.W. Walter U. Cell. 1994; 78: 919-925Abstract Full Text PDF PubMed Scopus (1495) Google Scholar, 3Knowles R.G. Moncada S. Biochem. J. 1994; 298: 249-258Crossref PubMed Scopus (2485) Google Scholar for reviews). NO is synthesized from arginine by nitric-oxide synthase (NOS), generating citrulline. Cellular NO production is absolutely dependent on the availability of arginine. This amino acid can be obtained from exogenous sources via the blood circulation, from intracellular protein degradation, or by the endogenous synthesis of arginine. Major sites of arginine synthesis in ureotelic animals are the liver, where arginine generated in the urea cycle (ornithine cycle) is rapidly converted to urea and ornithine by arginase, and the kidney, where arginine is synthesized from citrulline and released into the blood circulation (see Ref. 4Morris S.M. Annu. Rev. Nutr. 1994; 12: 81-101Crossref Scopus (214) Google Scholar for a review). In other tissues and cell types, arginine can be generated from citrulline, which is produced as a coproduct of the NOS reaction, forming a cycle that is composed of NOS, argininosuccinate synthetase, and argininosuccinate lyase and that is termed the “citrulline-NO cycle” (5Hecker M. Sessa W.C. Harris H.J. Ånggárd E.E. Vane J.R. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 8612-8616Crossref PubMed Scopus (403) Google Scholar, 6Mitchell J.A. Hecker M. Vane J.R. Eur. J. Pharmacol. 1990; 176: 253-254Crossref PubMed Scopus (48) Google Scholar, 7Wu G. Brosnan J.T. Biochem. J. 1992; 281: 45-48Crossref PubMed Scopus (161) Google Scholar, 8Nussler A.K. Billiar T.R. Liu Z.-Z. Morris Jr., S.M. J. Biol. Chem. 1994; 269: 1257-1261Abstract Full Text PDF PubMed Google Scholar, 9Hattori Y. Campbell E.B. Gross S.S. J. Biol. Chem. 1994; 269: 9405-9408Abstract Full Text PDF PubMed Google Scholar, 10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar). The inducible isoform of NOS (iNOS) and argininosuccinate synthetase are coinduced in activated murine macrophage-like RAW 264.7 cells (8Nussler A.K. Billiar T.R. Liu Z.-Z. Morris Jr., S.M. J. Biol. Chem. 1994; 269: 1257-1261Abstract Full Text PDF PubMed Google Scholar), in cultured vascular smooth muscle cells (9Hattori Y. Campbell E.B. Gross S.S. J. Biol. Chem. 1994; 269: 9405-9408Abstract Full Text PDF PubMed Google Scholar), and in vivo (10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar, 11Hattori Y. Shimoda S. Gross S.S. Biochem. Biophys. Res. Commun. 1995; 215: 148-153Crossref PubMed Scopus (28) Google Scholar). Argininosuccinate lyase is also induced in vivo (10Nagasaki A. Gotoh T. Takeya M. Yu Y. Takiguchi M. Matsuzaki H. Takatsuki K. Mori M. J. Biol. Chem. 1996; 271: 2658-2662Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar, 11Hattori Y. Shimoda S. Gross S.S. Biochem. Biophys. Res. Commun. 1995; 215: 148-153Crossref PubMed Scopus (28) Google Scholar).On the other hand, arginine is utilized for both the arginase and NOS reactions. Thus, these two enzymes compete for arginine. At least two isoforms of arginase are present. Liver-type arginase (arginase I) is expressed almost exclusively in the liver and catalyzes the final step of urea synthesis. Arginase I activity (12Lamers W.H. Mooren P.G. De Graaf A. Charles R. Eur. J. Biochem. 1985; 146: 475-480Crossref PubMed Scopus (42) Google Scholar) and mRNA (13Morris Jr., S.M. Kepka D.M. Sweeney Jr., W.E. Avner E.D. Arch. Biochem. Biophys. 1989; 269: 175-180Crossref PubMed Scopus (34) Google Scholar) in rat liver increase markedly in the perinatal period, in coordination with other urea cycle enzymes. The enzyme is regulated by dietary protein (14Schimke R.T. J. Biol. Chem. 1962; 237: 459-468Abstract Full Text PDF PubMed Google Scholar) and hormones (15Nebes V.L. Morris Jr., S.M. Mol. Endocrinol. 1988; 2: 444-451Crossref PubMed Scopus (89) Google Scholar). Arginase I consists of three identical subunits of about 35,000 Da. cDNA clones were isolated from rat (16Dizikes G.J. Spector E.B. Cederbaum S.D. Somatic Cell Mol. Genet. 1986; 12: 375-384Crossref PubMed Scopus (43) Google Scholar, 17Kawamoto S. Amaya Y. Murakami K. Tokunaga F. Iwanaga S. Kobayashi K. Saheki T. Kimura S. Mori M. J. Biol. Chem. 1987; 262: 6280-6283Abstract Full Text PDF PubMed Google Scholar) and human (18Dizikes G.J. Grody W.W. Kern R.M. Cederbaum S.D. Biochem. Biophys. Res. Commun. 1986; 141: 53-59Crossref PubMed Scopus (68) Google Scholar, 19Haraguchi Y. Takiguchi M. Amaya Y. Kawamoto S. Matsuda I. Mori M. Proc. Natl. Acad. Sci. U. S. A. 1987; 84: 412-415Crossref PubMed Scopus (114) Google Scholar) liver. The rat (20Ohtake A. Takiguchi M. Shigeto Y. Amaya Y. Kawamoto S. Mori M. J. Biol. Chem. 1988; 263: 2245-2249Abstract Full Text PDF PubMed Google Scholar) and human (21Takiguchi M. Haraguchi Y. Mori M. Nucleic Acids Res. 1988; 16: 8789-8802Crossref PubMed Scopus (49) Google Scholar) genes are 11.5-12 kb long and consist of 8 exons. Promoter and enhancer regions of the rat gene were characterized (22Takiguchi M. Mori M. J. Biol. Chem. 1991; 266: 9186-9193Abstract Full Text PDF PubMed Google Scholar, 23Gotoh T. Haraguchi Y. Takiguchi M. Mori M. J. Biochem. 1994; 115: 778-788Crossref PubMed Scopus (26) Google Scholar, 24Chowdhury S. Gotoh T. Mori M. Takiguchi M. Eur. J. Biochem. 1996; 236: 500-509Crossref PubMed Scopus (32) Google Scholar). In addition to arginase I, an isozyme (arginase II) is present in extrahepatic tissues, including kidney, small intestine, and lactating mammary gland (25Glass R.D. Knox W.E. J. Biol. Chem. 1973; 248: 5785-5789Abstract Full Text PDF PubMed Google Scholar, 26Kaysen G.A. Strecker H.J. Biochem. J. 1973; 133: 779-788Crossref PubMed Scopus (127) Google Scholar, 27Herzfeld A. Raper S.M. Biochem. J. 1976; 153: 469-478Crossref PubMed Scopus (144) Google Scholar). The coinduction of NOS and arginase II activities in RAW 264.7 cells activated by lipopolysaccharide (LPS) was reported (28Wang W.W. Jenkinson C.P. Griscavage J.M. Kern R.M. Arabolos N.S. Byrns R.E. Cederbaum S.D. Ignarro L.J. Biochem. Biophys. Res. Commun. 1995; 210: 1009-1016Crossref PubMed Scopus (204) Google Scholar). We isolated cDNA for arginase II and showed that it is 59% identical with arginase I on the amino acid level (29Gotoh T. Sonoki T. Nagasaki A. Terada K. Takiguchi M. Mori M. FEBS Lett. 1996; 395: 119-122Crossref PubMed Scopus (210) Google Scholar). We also found that the enzyme is mitochondrial and that its mRNA is induced by LPS, dexamethasone, and cyclic adenosine monophosphate and is reduced by interferon-γ (29Gotoh T. Sonoki T. Nagasaki A. Terada K. Takiguchi M. Mori M. FEBS Lett. 1996; 395: 119-122Crossref PubMed Scopus (210) Google Scholar). To better understand the role of arginase isoforms in NO synthesis, we examined expression of the isoforms and iNOS in cultured rat peritoneal cells and in LPS-treated rats using RNA blots, immunoblots, and immunocytochemical analyses. We report here that iNOS and arginase I mRNAs and proteins are coinduced by LPS in cultured rat peritoneal macrophages and in the lung and spleen in vivo The induction of mRNA for CCAAT/enhancer-binding protein β (C/EBPβ), a transactivator of the arginase I gene, is also described.