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Satoru Takase,
Jun-Ichi Osuga,
Hayato Fujita,
Kazuo Hara, Motohiro Sekiya,
Masaki Igarashi,
Mikio Takanashi,
Yoshinori Takeuchi,
Yoshihiko Izumida,
Keisuke Ohta, [......],
Yoko Iizuka,
Ken Ohashi,
Hiroshi Yoshida,
Hidekatsu Yanai,
Norio Tada,
Takanari Gotoda,
Shun Ishibashi,
Takashi Kadowaki,
Hiroaki Okazaki,
Naoya Yahagi
[show abstract]
[hide abstract]
ABSTRACT: Aim: Familial apolipoprotein C-II (apoC-II) deficiency is a rare autosomal recessive disorder with marked hypertriglyceridemia resulting from impaired activation of lipoprotein lipase. In most cases of apoC-II deficiency, causative mutations have been found in the protein-coding region of APOC2; however, several atypical cases of apoC-II deficiency were reported to have markedly reduced, but detectable levels of plasma apoC-II protein (hereafter referred to as hypoapoC-II), which resulted from decreased promoter activity or improper splicing of apoC-II mRNA due to homozygous mutations in APOC2. Here we aim to dissect the molecular bases of a new case of hypoapoC-II.Methods: We performed detailed biochemical/genetic analyses of our new case of hypoapoC-II, manifesting severe hypertriglyceridemia (plasma triglycerides, 3235 mg·dL-1) with markedly reduced levels of plasma apoC-II (0.6 mg·dL-1).Results: We took advantage of a monocyte/macrophage culture system to prove that transcription of apoC-II mRNA was decreased in the patient's cells, which is compatible with the reported features of hypoapoC-II. Concomitantly, transcriptional activity of the minigene reporter construct of the patient's APOC2 gene was decreased; however, no rare variant was detected in the patient's APOC2 gene. Fifty single nucleotide variants were detected in the patient's APOC2, but all were common variants (allele frequencies ＞35%) that are supposedly not causative.Conclusions: A case of apoC-II deficiency was found that is phenotypically identical to hypoapoC-II but with no causative mutations in APOC2, implying that other genes regulate apoC-II levels. The clinical entity of hypoapoC-II is discussed.
Journal of atherosclerosis and thrombosis 03/2013; · 2.69 Impact Factor
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Shuichi Nagashima,
Hiroaki Yagyu,
Ken Ohashi,
Fumiko Tazoe,
Manabu Takahashi,
Taichi Ohshiro,
Tumenbayar Bayasgalan,
Kenta Okada, Motohiro Sekiya,
Jun-ichi Osuga,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) catalyzes the rate-limiting step in cholesterol biosynthesis and has proven to be an effective target of lipid-lowering drugs, statins. The aim of this study was to understand the role of hepatic HMGCR in vivo.
To disrupt the HMGCR gene in liver, we generated mice homozygous for a floxed HMGCR allele and heterozygous for a transgene encoding Cre recombinase under the control of the albumin promoter (liver-specific HMGCR knockout mice). Ninety-six percent of male and 71% of female mice died by 6 weeks of age, probably as a result of liver failure or hypoglycemia. At 5 weeks of age, liver-specific HMGCR knockout mice showed severe hepatic steatosis with apoptotic cells, hypercholesterolemia, and hypoglycemia. The hepatic steatosis and death were completely reversed by providing the animals with mevalonate, indicating its essential role in normal liver function. There was a modest decrease in hepatic cholesterol synthesis in liver-specific HMGCR knockout mice. Instead, they showed a robust increase in the fatty acid synthesis, independent of sterol regulatory element binding protein-1c.
Hepatocyte HMGCR is essential for the survival of mice, and its abrogation elicits hepatic steatosis with jaundice and hypoglycemia.
Arteriosclerosis Thrombosis and Vascular Biology 06/2012; 32(8):1824-31. · 6.37 Impact Factor
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Ryo Saito,
Takashi Matsuzaka,
Tadayoshi Karasawa, Motohiro Sekiya,
Nazuki Okada,
Masaki Igarashi,
Rie Matsumori,
Kiyoaki Ishii,
Yoshimi Nakagawa,
Hitoshi Iwasaki,
Kazuto Kobayashi,
Shigeru Yatoh,
Akimitsu Takahashi,
Hirohito Sone,
Hiroaki Suzuki,
Naoya Yahagi,
Nobuhiro Yamada,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Elovl6, a long-chain fatty acid elongase, is a rate-limiting enzyme that elongates saturated and monounsaturated fatty acids and has been shown to be related to obesity-induced insulin resistance via modification of fatty acid composition. In this study, we investigated the roles of Elovl6 in foam cell formation in macrophages and atherosclerosis in mice.
To investigate the roles of Elovl6 in macrophages in the progression of atherosclerosis, we transplanted bone marrow cells of wild-type or Elovl6(-/-) mice into irradiated LDL-R(-/-) mice that were fed a western diet. Aortic atherosclerotic lesion areas and infiltration of macrophages were significantly smaller in Elovl6(-/-) bone marrow cells-transplanted LDL-R(-/-) mice than in wild-type. Accumulation of esterified cholesterol on exposure to acetylated-LDL was less severe in peritoneal macrophages from Elovl6(-/-) mice than those from wild-type. Cholesterol efflux and expression of cholesterol efflux transporters were increased in Elovl6(-/-) macrophages, although no difference in uptake of acetylated-LDL was found between the two groups. On analysis of fatty acid composition of the esterified cholesterol fraction in macrophages, n-6 polyunsaturated fatty acids were decreased by absence of Elovl6.
These findings suggest that Elovl6 in macrophages may contribute to foam cell formation and progression of atherosclerosis.
Arteriosclerosis Thrombosis and Vascular Biology 09/2011; 31(9):1973-9. · 6.37 Impact Factor
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Tadayoshi Karasawa,
Akimitsu Takahashi,
Ryo Saito, Motohiro Sekiya,
Masaki Igarashi,
Hitoshi Iwasaki,
Shoko Miyahara,
Saori Koyasu,
Yoshimi Nakagawa,
Kiyoaki Ishii,
Takashi Matsuzaka,
Kazuto Kobayashi,
Naoya Yahagi,
Kazuhiro Takekoshi,
Hirohito Sone,
Shigeru Yatoh,
Hiroaki Suzuki,
Nobuhiro Yamada,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Sterol regulatory element-binding protein-1 (SREBP-1) is nutritionally regulated and is known to be a key transcription factor regulating lipogenic enzymes. The goal of this study was to evaluate the roles of SREBP-1 in dyslipidemia and atherosclerosis.
Transgenic mice that overexpress SREBP-1c in the liver and SREBP-1-deficient mice were crossed with low-density lipoprotein receptor (LDLR)-deficient mice, and the plasma lipids and atherosclerosis were analyzed. Hepatic SREBP-1c overexpression in LDLR-deficient mice caused postprandial hypertriglyceridemia, increased very-low-density lipoprotein (VLDL) cholesterol, and decreased high-density lipoprotein cholesterol in plasma, which resulted in accelerated aortic atheroma formation. Conversely, absence of SREBP-1 suppressed Western diet-induced hyperlipidemia in LDLR-deficient mice and ameliorated atherosclerosis. In contrast, bone marrow-specific SREBP-1 deficiency did not alter the development of atherosclerosis. The size of nascent VLDL particles secreted from the liver was increased in SREBP-1c transgenic mice and reduced in SREBP-1-deficient mice, accompanied by upregulation and downregulation of phospholipid transfer protein expression, respectively.
Hepatic SREBP-1c determines plasma triglycerides and remnant cholesterol and contributes to atherosclerosis in hyperlipidemic states. Hepatic SREBP-1c also regulates the size of nascent VLDL particles.
Arteriosclerosis Thrombosis and Vascular Biology 05/2011; 31(8):1788-95. · 6.37 Impact Factor
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[show abstract]
[hide abstract]
ABSTRACT: Cholesterol ester-laden macrophage foam cells are a hallmark of atherosclerosis. The cycle of esterification and hydrolysis of cholesterol esters is one of the key steps in macrophage cholesterol trafficking. In the process of foam cell formation, excess free cholesterol undergoes esterification by acyl coenzyme A: acylcholesterol transferase 1 (ACAT-1), and fatty acid sterol esters are stored in cytoplasmic lipid droplets. The actions of ACAT-1 are opposed by neutral cholesterol ester hydrolase (nCEH), which generates free cholesterol and fatty acids. The resulting free cholesterol is a preferential source for cholesterol efflux into the extracellular space. Despite the important role of nCEH in protection against foam cell formation and atherosclerosis, the molecular identity of nCEH has long been debated. Although hormone-sensitive lipase (LIPE) has been proposed to be the nCEH in macrophages, recent evidence suggested the existence of other nCEH(s). We have recently identified a novel nCEH, neutral cholesterol ester hydrolase 1 (NCEH1), and demonstrated that NCEH1, in addition to LIPE, primarily mediates the hydrolysis of CE in macrophages. This review focuses on the protective roles of nCEHs in atherosclerosis, with special emphasis on the role of NCEH1.
Journal of atherosclerosis and thrombosis 04/2011; 18(5):359-64. · 2.69 Impact Factor
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Keisuke Ohta, Motohiro Sekiya,
Hiroshi Uozaki,
Masaki Igarashi,
Satoru Takase,
Masayoshi Kumagai,
Mikio Takanashi,
Yoshinori Takeuchi,
Yoshihiko Izumida,
Midori Kubota,
Makiko Nishi,
Hiroaki Okazaki,
Yoko Iizuka,
Naoya Yahagi,
Hiroaki Yagyu,
Masashi Fukayama,
Takashi Kadowaki,
Ken Ohashi,
Shun Ishibashi,
Jun-ichi Osuga
[show abstract]
[hide abstract]
ABSTRACT: We have previously demonstrated that neutral cholesterol ester hydrolase 1 (Nceh1) regulates foam cell formation and atherogenesis through the catalytic activity of cholesterol ester hydrolysis, and that Nceh1 and hormone-sensitive lipase (Lipe) are responsible for the majority of neutral cholesterol ester hydrolase activity in macrophages. There are several cholesterol ester-metabolizing tissues and cells other than macrophages, among which adrenocortical cells are also known to utilize the intracellular cholesterol for steroidogenesis. It has been believed that the mobilization of intracellular cholesterol ester in adrenal glands was facilitated solely by Lipe. We herein demonstrate that Nceh1 is also involved in cholesterol ester hydrolysis in adrenal glands. While Lipe deficiency remarkably reduced the neutral cholesterol ester hydrolase activity in adrenal glands as previously reported, additional inactivation of Nceh1 gene completely abrogated the activity. Adrenal glands were enlarged in proportion to the degree of reduced neutral cholesterol ester hydrolase activity, and the enlargement of adrenal glands and the accumulation of cholesterol esters were most pronounced in the Nceh1/Lipe double-deficient mice. Thus Nceh1 is involved in the adrenal cholesterol metabolism, and the cholesterol ester hydrolytic activity in adrenal glands is associated with the organ enlargement.
Biochemical and Biophysical Research Communications 01/2011; 404(1):254-60. · 2.48 Impact Factor
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Masaki Igarashi,
Jun-ichi Osuga,
Hiroshi Uozaki, Motohiro Sekiya,
Shuichi Nagashima,
Manabu Takahashi,
Satoru Takase,
Mikio Takanashi,
Yongxue Li,
Keisuke Ohta, [......],
Makiko Nishi,
Masakiyo Hosokawa,
Christian Fledelius,
Poul Jacobsen,
Hiroaki Yagyu,
Masashi Fukayama,
Ryozo Nagai,
Takashi Kadowaki,
Ken Ohashi,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: Hydrolysis of intracellular cholesterol ester (CE) is the key step in the reverse cholesterol transport in macrophage foam cells. We have recently shown that neutral cholesterol ester hydrolase (Nceh)1 and hormone-sensitive lipase (Lipe) are key regulators of this process in mouse macrophages. However, it remains unknown which enzyme is critical in human macrophages and atherosclerosis.
We aimed to identify the enzyme responsible for the CE hydrolysis in human macrophages and to determine its expression in human atherosclerosis.
We compared the expression of NCEH1, LIPE, and cholesterol ester hydrolase (CES1) in human monocyte-derived macrophages (HMMs) and examined the effects of inhibition or overexpression of each enzyme in the cholesterol trafficking. The pattern of expression of NCEH1 was similar to that of neutral CE hydrolase activity during the differentiation of HMMs. Overexpression of human NCEH1 increased the hydrolysis of CE, thereby stimulating cholesterol mobilization from THP-1 macrophages. Knockdown of NCEH1 specifically reduced the neutral CE hydrolase activity. Pharmacological inhibition of NCEH1 also increased the cellular CE in HMMs. In contrast, LIPE was barely detectable in HMMs, and its inhibition did not decrease neutral CE hydrolase activity. Neither overexpression nor knockdown of CES1 affected the neutral CE hydrolase activity. NCEH1 was expressed in CD68-positive macrophage foam cells of human atherosclerotic lesions.
NCEH1 is expressed in human atheromatous lesions, where it plays a critical role in the hydrolysis of CE in human macrophage foam cells, thereby contributing to the initial part of reverse cholesterol transport in human atherosclerosis.
Circulation Research 11/2010; 107(11):1387-95. · 9.49 Impact Factor
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Yoshinori Takeuchi,
Naoya Yahagi,
Yoshihiko Izumida,
Makiko Nishi,
Midori Kubota,
Yuji Teraoka,
Takashi Yamamoto,
Takashi Matsuzaka,
Yoshimi Nakagawa, Motohiro Sekiya, [......],
Ken Ohashi,
Jun-ichi Osuga,
Takanari Gotoda,
Shun Ishibashi,
Keiji Itaka,
Kazunori Kataoka,
Ryozo Nagai,
Nobuhiro Yamada,
Takashi Kadowaki,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Sterol regulatory element-binding protein (SREBP)-1 is a key transcription factor for the regulation of lipogenic enzyme genes
in the liver. Polyunsaturated fatty acids (PUFA) selectively suppress hepatic SREBP-1, but molecular mechanisms remain largely
unknown. To gain insight into this regulation, we established in vivo reporter assays to assess the activities of Srebf1c transcription and proteolytic processing. Using these in vivo reporter assays, we showed that the primary mechanism for PUFA suppression of SREBP-1 is at the proteolytic processing level
and that this suppression in turn decreases the mRNA transcription through lowering SREBP-1 binding to the SREBP-binding element
on the promoter (“autoloop regulatory circuit”), although liver X receptor, an activator for Srebf1c transcription, is not involved in this regulation by PUFA. The mechanisms for PUFA suppression of SREBP-1 confirm that the
autoloop regulation for transcription is crucial for the nutritional regulation of triglyceride synthesis.
Journal of Biological Chemistry 04/2010; 285(15):11681-11691. · 4.77 Impact Factor
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Yoshinori Takeuchi,
Naoya Yahagi,
Yoshihiko Izumida,
Makiko Nishi,
Midori Kubota,
Yuji Teraoka,
Takashi Yamamoto,
Takashi Matsuzaka,
Yoshimi Nakagawa, Motohiro Sekiya, [......],
Ken Ohashi,
Jun-ichi Osuga,
Takanari Gotoda,
Shun Ishibashi,
Keiji Itaka,
Kazunori Kataoka,
Ryozo Nagai,
Nobuhiro Yamada,
Takashi Kadowaki,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Sterol regulatory element-binding protein (SREBP)-1 is a key transcription factor for the regulation of lipogenic enzyme genes in the liver. Polyunsaturated fatty acids (PUFA) selectively suppress hepatic SREBP-1, but molecular mechanisms remain largely unknown. To gain insight into this regulation, we established in vivo reporter assays to assess the activities of Srebf1c transcription and proteolytic processing. Using these in vivo reporter assays, we showed that the primary mechanism for PUFA suppression of SREBP-1 is at the proteolytic processing level and that this suppression in turn decreases the mRNA transcription through lowering SREBP-1 binding to the SREBP-binding element on the promoter ("autoloop regulatory circuit"), although liver X receptor, an activator for Srebf1c transcription, is not involved in this regulation by PUFA. The mechanisms for PUFA suppression of SREBP-1 confirm that the autoloop regulation for transcription is crucial for the nutritional regulation of triglyceride synthesis.
Journal of Biological Chemistry 02/2010; 285(15):11681-91. · 4.77 Impact Factor
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Motohiro Sekiya,
Jun-Ichi Osuga,
Shuichi Nagashima,
Taichi Ohshiro,
Masaki Igarashi,
Hiroaki Okazaki,
Manabu Takahashi,
Fumiko Tazoe,
Taeko Wada,
Keisuke Ohta, [......],
Masayoshi Kumagai,
Makiko Nishi,
Satoru Takase,
Naoya Yahagi,
Hiroaki Yagyu,
Ken Ohashi,
Ryozo Nagai,
Takashi Kadowaki,
Yusuke Furukawa,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: Cholesterol ester (CE)-laden macrophage foam cells are the hallmark of atherosclerosis, and the hydrolysis of intracellular CE is one of the key steps in foam cell formation. Although hormone-sensitive lipase (LIPE) and cholesterol ester hydrolase (CEH), which is identical to carboxylsterase 1 (CES1, hCE1), were proposed to mediate the neutral CE hydrolase (nCEH) activity in macrophages, recent evidences have suggested the involvement of other enzymes. We have recently reported the identification of a candidate, neutral cholesterol ester hydrolase 1(Nceh1). Here we demonstrate that genetic ablation of Nceh1 promotes foam cell formation and the development of atherosclerosis in mice. We further demonstrate that Nceh1 and Lipe mediate a comparable degree of nCEH activity in macrophages and together account for most of the activity. Mice lacking both Nceh1 and Lipe aggravated atherosclerosis in an additive manner. Thus, Nceh1 is a promising target for the treatment of atherosclerosis.
Cell metabolism 10/2009; 10(3):219-28. · 17.35 Impact Factor
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Motohiro Sekiya,
Naoya Yahagi,
Yoshiaki Tamura,
Hiroaki Okazaki,
Masaki Igarashi,
Keisuke Ohta,
Mikio Takanashi,
Masayoshi Kumagai,
Satoru Takase,
Makiko Nishi, [......],
Ken Ohashi,
Yoko Iizuka,
Hiroaki Yagyu,
Takanari Gotoda,
Ryozo Nagai,
Hitoshi Shimano,
Nobuhiro Yamada,
Takashi Kadowaki,
Shun Ishibashi,
Jun-ichi Osuga
[show abstract]
[hide abstract]
ABSTRACT: It has long been a matter of debate whether the hormone-sensitive lipase (HSL)-mediated lipolysis in pancreatic beta-cells can affect insulin secretion through the alteration of lipotoxicity. We generated mice lacking both leptin and HSL Lep(ob/ob)/HSL(-/-) and explored the role of HSL in pancreatic beta-cells in the setting of obesity. Lep(ob/ob)/HSL(-/-) developed elevated blood glucose levels and reduced plasma insulin levels compared with Lep(ob/ob)/HSL(+/+) in a fed state, while the deficiency of HSL did not affect glucose homeostasis in Lep(+/+) background. The deficiency of HSL exacerbated the accumulation of triglycerides in Lep(ob/ob) islets, leading to reduced glucose-stimulated insulin secretion. The deficiency of HSL also diminished the islet mass in Lep(ob/ob) mice due to decreased cell proliferation. In conclusion, HSL affects insulin secretary capacity especially in the setting of obesity.
Biochemical and Biophysical Research Communications 08/2009; 387(3):511-5. · 2.48 Impact Factor
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Masaki Igarashi,
Jun-Ichi Osuga,
Masashi Isshiki, Motohiro Sekiya,
Hiroaki Okazaki,
Satoru Takase,
Mikio Takanashi,
Keisuke Ohta,
Masayoshi Kumagai,
Makiko Nishi,
Toshiro Fujita,
Ryozo Nagai,
Takashi Kadowaki,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: Neutral cholesterol ester hydrolase (NCEH) accounts for a large part of the nCEH activity in macrophage foam cells, a hallmark of atherosclerosis, but its subcellular localization and structure-function relationship are unknown. Here, we determined subcellular localization, glycosylation, and nCEH activity of a series of NCEH mutants expressed in macrophages. NCEH is a single-membrane-spanning type II membrane protein comprising three domains: N-terminal, catalytic, and lipid-binding domains. The N-terminal domain serves as a type II signal anchor sequence to recruit NCEH to the endoplasmic reticulum (ER) with its catalytic domain within the lumen. All of the putative N-linked glycosylation sites (Asn(270), Asn(367), and Asn(389)) of NCEH are glycosylated. Glycosylation at Asn(270), which is located closest to the catalytic serine motif, is important for the enzymatic activity. Cholesterol loading by incubation with acetyl-LDL does not change the ER localization of NCEH. In conclusion, NCEH is targeted to the ER of macrophages, where it hydrolyzes CE to deliver cholesterol for efflux out of the cells.
The Journal of Lipid Research 08/2009; 51(2):274-85. · 5.56 Impact Factor
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Hiroaki Okazaki,
Masaki Igarashi,
Makiko Nishi, Motohiro Sekiya,
Makiko Tajima,
Satoru Takase,
Mikio Takanashi,
Keisuke Ohta,
Yoshiaki Tamura,
Sachiko Okazaki,
Naoya Yahagi,
Ken Ohashi,
Michiyo Amemiya-Kudo,
Yoshimi Nakagawa,
Ryozo Nagai,
Takashi Kadowaki,
Jun-ichi Osuga,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: Unstable lipid-rich plaques in atherosclerosis are characterized by the accumulation of macrophage foam cells loaded with cholesterol ester (CE). Although hormone-sensitive lipase and cholesteryl ester hydrolase (CEH) have been proposed to mediate the hydrolysis of CE in macrophages, circumstantial evidence suggests the presence of other enzymes with neutral cholesterol ester hydrolase (nCEH) activity. Here we show that the murine orthologue of KIAA1363, designated as neutral cholesterol ester hydrolase (NCEH), is a microsomal nCEH with high expression in murine and human macrophages. The effect of various concentrations of NaCl on its nCEH activity resembles that on endogenous nCEH activity of macrophages. RNA silencing of NCEH decreases nCEH activity at least by 50%; conversely, its overexpression inhibits the CE formation in macrophages. Immunohistochemistry reveals that NCEH is expressed in macrophage foam cells in atherosclerotic lesions. These data indicate that NCEH is responsible for a major part of nCEH activity in macrophages and may be a potential therapeutic target for the prevention of atherosclerosis.
Journal of Biological Chemistry 10/2008; 283(48):33357-64. · 4.77 Impact Factor
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Motohiro Sekiya,
Jun-Ichi Osuga,
Naoya Yahagi,
Hiroaki Okazaki,
Yoshiaki Tamura,
Masaki Igarashi,
Satoru Takase,
Kenji Harada,
Sachiko Okazaki,
Yoko Iizuka,
Ken Ohashi,
Hiroaki Yagyu,
Mitsuyo Okazaki,
Takanari Gotoda,
Ryozo Nagai,
Takashi Kadowaki,
Hitoshi Shimano,
Nobuhiro Yamada,
Shun Ishibashi
[show abstract]
[hide abstract]
ABSTRACT: Hormone-sensitive lipase (HSL) regulates the hydrolysis of acylglycerol and cholesteryl ester (CE) in various organs, including adipose tissues. However, the hepatic expression level of HSL has been reported to be almost negligible. In the present study, we found that mice lacking both leptin and HSL (Lep(ob/ob)/HSL(-/-)) showed massive accumulation of CE in the liver compared with Lep(ob/ob)/HSL(+/+) mice, while triacylglycerol (TG) accumulation was modest. Similarly, feeding with a high-cholesterol diet induced hepatic CE accumulation in HSL(-/-) mice. Supporting these observations, we detected significant expression of protein as well as mRNA of HSL in the liver. HSL(-/-) mice showed reduced activity of CE hydrolase, but not of TG lipase, in the liver compared with wild-type mice. Furthermore, we confirmed the expression of HSL in viable parenchymal cells isolated from wild-type mice. The hepatocytes from HSL(-/-) mice showed reduced activity of CE hydrolase and contained more CE than those from HSL(+/+) mice even without the incubation with lipoproteins. Incubation with LDL further augmented the accumulation of CE in the HSL-deficient hepatocytes. From these results, we conclude that HSL is involved in the hydrolysis of CE in hepatocyes.
The Journal of Lipid Research 09/2008; 49(8):1829-38. · 5.56 Impact Factor
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Yoshinori Takeuchi,
Naoya Yahagi,
Yoshimi Nakagawa,
Takashi Matsuzaka,
Ritsuko Shimizu, Motohiro Sekiya,
Yoko Iizuka,
Ken Ohashi,
Takanari Gotoda,
Masayuki Yamamoto,
Ryozo Nagai,
Takashi Kadowaki,
Nobuhiro Yamada,
Jun-Ichi Osuga,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Sterol regulatory element-binding protein (SREBP)-1c is the master regulator of lipogenic gene expression in liver. The mRNA abundance of SREBP-1c is markedly induced when animals are refed after starvation, although the regulatory mechanism is so far unknown. To investigate the mechanism of refeeding response of SREBP-1c gene expression in vivo, we generated a transgenic mouse model that carries 2.2kb promoter region fused to the luciferase reporter gene. These transgenic mice exhibited refeeding responses of the reporter in liver and adipose tissues with extents essentially identical to those of endogenous SREBP-1c mRNA. The same results were obtained from experiments using adenovirus-mediated SREBP-1c-promoter-luciferase fusion gene transduction to liver. These data demonstrate that the regulation of SREBP-1c gene expression is at the transcription level, and that the 2.2kb 5'-flanking region is sufficient for this regulation. Moreover, when these transgenic or adenovirus-infected mice were placed on insulin-depleted state by streptozotocin treatment, the reporter expression was upregulated as strongly as in control mice, demonstrating that this regulation is not dominated by serum insulin level. These mice are the first models to provide the mechanistic insight into the transcriptional regulation of SREBP-1c gene in vivo.
Biochemical and Biophysical Research Communications 12/2007; 363(2):329-35. · 2.48 Impact Factor
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Takashi Matsuzaka,
Hitoshi Shimano,
Naoya Yahagi,
Toyonori Kato,
Ayaka Atsumi,
Takashi Yamamoto,
Noriyuki Inoue,
Mayumi Ishikawa,
Sumiyo Okada,
Naomi Ishigaki, [......],
Ken Ohashi,
Alyssa H Hasty,
Yoshimi Nakagawa,
Akimitsu Takahashi,
Hiroaki Suzuki,
Sigeru Yatoh,
Hirohito Sone,
Hideo Toyoshima,
Jun-ichi Osuga,
Nobuhiro Yamada
[show abstract]
[hide abstract]
ABSTRACT: Insulin resistance is often associated with obesity and can precipitate type 2 diabetes. To date, most known approaches that improve insulin resistance must be preceded by the amelioration of obesity and hepatosteatosis. Here, we show that this provision is not mandatory; insulin resistance and hyperglycemia are improved by the modification of hepatic fatty acid composition, even in the presence of persistent obesity and hepatosteatosis. Mice deficient for Elovl6, the gene encoding the elongase that catalyzes the conversion of palmitate to stearate, were generated and shown to become obese and develop hepatosteatosis when fed a high-fat diet or mated to leptin-deficient ob/ob mice. However, they showed marked protection from hyperinsulinemia, hyperglycemia and hyperleptinemia. Amelioration of insulin resistance was associated with restoration of hepatic insulin receptor substrate-2 and suppression of hepatic protein kinase C epsilon activity resulting in restoration of Akt phosphorylation. Collectively, these data show that hepatic fatty acid composition is a new determinant for insulin sensitivity that acts independently of cellular energy balance and stress. Inhibition of this elongase could be a new therapeutic approach for ameliorating insulin resistance, diabetes and cardiovascular risks, even in the presence of a continuing state of obesity.
Nature Medicine 11/2007; 13(10):1193-202. · 22.46 Impact Factor
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Motohiro Sekiya,
Naoya Yahagi,
Takashi Matsuzaka,
Yoshinori Takeuchi,
Yoshimi Nakagawa,
Haruka Takahashi,
Hiroaki Okazaki,
Yoko Iizuka,
Ken Ohashi,
Takanari Gotoda,
Shun Ishibashi,
Ryozo Nagai,
Tsutomu Yamazaki,
Takashi Kadowaki,
Nobuhiro Yamada,
Jun-ichi Osuga,
Hitoshi Shimano
[show abstract]
[hide abstract]
ABSTRACT: Sterol regulatory element-binding protein (SREBP)-1c is now well established as a key transcription factor for the regulation of lipogenic enzyme genes such as FAS in hepatocytes. Meanwhile, the mechanisms of lipogenic gene regulation in adipocytes remain unclear. Here, we demonstrate that those in adipocytes are independent of SREBP-1c. In adipocytes, unlike in hepatocytes, the stimulation of SREBP-1c expression by liver X receptor agonist does not accompany lipogenic gene upregulation, although nuclear SREBP-1c protein is concomitantly increased, indicating that the activation process of SREBP-1c by the cleavage system is intact in adipocytes. Supportively, transcriptional activity of the mature form of SREBP-1c for the FAS promoter was negligible when measured by reporter analysis. As an underlying mechanism, accessibility of SREBP-1c to the functional elements was involved, because chromatin immunoprecipitation assays revealed that SREBP-1c does not bind to the functional SRE/E-box site on the FAS promoter in adipocytes. Moreover, genetic disruption of SREBP-1 did not cause any changes in lipogenic gene expression in adipose tissue. In summary, in adipocytes, unlike in hepatocytes, increments in nuclear SREBP-1c are not accompanied by transactivation of lipogenic genes; thus, SREBP-1c is not committed to the regulation of lipogenesis.
The Journal of Lipid Research 08/2007; 48(7):1581-91. · 5.56 Impact Factor
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Hiroaki Okazaki,
Fumiko Tazoe,
Sachiko Okazaki,
Naoyuki Isoo,
Kazuhisa Tsukamoto, Motohiro Sekiya,
Naoya Yahagi,
Yoko Iizuka,
Ken Ohashi,
Tetsuya Kitamine, [......],
Toshihiro Inaba,
Hiroaki Yagyu,
Mitsuyo Okazaki,
Hitoshi Shimano,
Norihito Shibata,
Hiroyuki Arai,
Ryo-Zo Nagai,
Takashi Kadowaki,
Jun-Ichi Osuga,
Shun Ishibashi
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ABSTRACT: Squalene synthase (SS) is the first committed enzyme for cholesterol biosynthesis, located at a branch point in the mevalonate pathway. To examine the role of SS in the overall cholesterol metabolism, we transiently overexpressed mouse SS in the livers of mice using adenovirus-mediated gene transfer. Overexpression of SS increased de novo cholesterol biosynthesis with increased 3-hydroxy-3-methyglutaryl-CoA (HMG-CoA) reductase activity, in spite of the downregulation of its own mRNA expression. Furthermore, overexpression of SS increased plasma concentrations of LDL, irrespective of the presence of functional LDL receptor (LDLR). Thus, the hypercholesterolemia is primarily caused by increased hepatic production of cholesterol-rich VLDL, as demonstrated by the increases in plasma cholesterol levels after intravenous injection of Triton WR1339. mRNA expression of LDLR was decreased, suggesting that defective LDL clearance contributed to the development of hypercholesterolemia. Curiously, the liver was enlarged, with a larger number of Ki-67-positive cells. These results demonstrate that transient upregulation of SS stimulates cholesterol biosynthesis as well as lipoprotein production, providing the first in vivo evidence that SS plays a regulatory role in cholesterol metabolism through modulation of HMG-CoA reductase activity and cholesterol biosynthesis.
The Journal of Lipid Research 10/2006; 47(9):1950-8. · 5.56 Impact Factor
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Hiroaki Okazaki,
Masaki Igarashi,
Makiko Nishi,
Makiko Tajima, Motohiro Sekiya,
Sachiko Okazaki,
Naoya Yahagi,
Ken Ohashi,
Kazuhisa Tsukamoto,
Michiyo Amemiya-Kudo, [......],
Hitoshi Shimano,
Nobuhiro Yamada,
Junken Aoki,
Rei Morikawa,
Yasukazu Takanezawa,
Hiroyuki Arai,
Ryozo Nagai,
Takashi Kadowaki,
Jun-Ichi Osuga,
Shun Ishibashi
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ABSTRACT: Molecular mechanisms underlying lipolysis, as defined by mobilization of fatty acids from adipose tissue, are not fully understood. A database search for enzymes with alpha/beta hydrolase folds, the GXSXG motif for serine esterase and the His-Gly dipeptide motif, has provided a previously unannotated gene that is induced during 3T3-L1 adipocytic differentiation. Because of its remarkable structural resemblance to triacylglycerol hydrolase (TGH) with 70.4% identity, we have tentatively designated this enzyme as TGH-2 and the original TGH as TGH-1. TGH-2 is also similar to TGH-1 in terms of tissue distribution, subcellular localization, substrate specificity, and regulation. Both enzymes are predominantly expressed in liver, adipose tissue, and kidney. In adipocytes, they are localized in microsome and fatcake. Both enzymes hydrolyzed p-nitophenyl butyrate, triolein, and monoolein but not diolein, cholesteryl oleate, or phospholipids; hydrolysis of short-chain fatty acid ester was 30,000-fold more efficient than that of long-chain fatty acid triacylglycerol. Fasting increased the expression of both genes in white adipose tissue, whereas refeeding suppressed their expression. RNA silencing of TGH-2 reduced isoproterenol-stimulated glycerol release by 10% in 3T3-L1 adipocytes, while its overexpression increased the glycerol release by 20%. Thus, TGH-2 may make a contribution to adipocyte lipolysis during period of increased energy demand.
Diabetes 08/2006; 55(7):2091-7. · 8.29 Impact Factor
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Yuho Najima,
Naoya Yahagi,
Yoshinori Takeuchi,
Takashi Matsuzaka, Motohiro Sekiya,
Yoshimi Nakagawa,
Michiyo Amemiya-Kudo,
Hiroaki Okazaki,
Sachiko Okazaki,
Yoshiaki Tamura,
Yoko Iizuka,
Ken Ohashi,
Kenji Harada,
Takanari Gotoda,
Ryozo Nagai,
Takashi Kadowaki,
Shun Ishibashi,
Nobuhiro Yamada,
Jun-ichi Osuga,
Hitoshi Shimano
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ABSTRACT: Sterol regulatory element-binding proteins (SREBPs) are transcription factors that are predominately involved in the regulation of lipogenic and cholesterogenic enzyme gene expression. To identify unknown proteins that interact with SREBP, we screened nuclear extract proteins with 35S-labeled SREBP-1 bait in Far Western blotting analysis. Using this approach, high mobility group protein-B1 (HMGB1), a chromosomal protein, was identified as a novel SREBP interacting protein. In vitro glutathione S-transferase pull-down and in vivo coimmunoprecipitation studies confirmed an interaction between HMGB1 and both SREBP-1 and -2. The protein-protein interaction was mediated through the helix-loop-helix domain of SREBP-1, residues 309-344, and the A box of HMGB1. Furthermore, an electrophoretic mobility shift assay demonstrated that HMGB1 enhances SREBPs binding to their cognate DNA sequences. Moreover, luciferase reporter analyses, including RNA interference technique showed that HMGB1 potentiates the transcriptional activities of SREBP in cultured cells. These findings raise the intriguing possibility that HMGB1 is potentially involved in the regulation of lipogenic and cholesterogenic gene transcription.
Journal of Biological Chemistry 08/2005; 280(30):27523-32. · 4.77 Impact Factor
