永井 良三

Expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1alpha (HIF-1alpha), a transcription factor involved in the VEGF induction, was examined in the myocardium. In the 10-day-old chick embryonic hearts and ventricular myocytes, the mRNA species encoding VEGF and HIF-1alpha were abundantly expressed even at the basal conditions. The steady-state levels of the VEGF mRNA were regulated by various protein kinases. In the cultured neonatal rat ventricular myocytes,VEGF expression was upregulated by lipopolysaccharide. These data suggest that VEGF and HIF-1alpha expressed in cardiac myocytes may play significant roles in cardiovascular development. Furthermore, the cardiac myocyte-derived VEGF may also contribute to the pathogenesis of systemic inflammatory response syndrome, including myocardial interstitial edema.

The background cationic current and the effects of lysophosphatidylcholine (LPC) were investigated in cultured rabbit coi-onary arterial smooth muscle cells (CASMCs) by using patch clamp techniques and [Ca2+](i) measurements. Here, we provided evidence that background nonselective cation current (I-NSC) contributed to form membrane potential, and LPC further induced I NSC' followed by depolarizing the membrane. The background and LPC-activated I-NSC were blocked by La3+ and Gd3+ but not by nicardipine and verapamil. LPC increased [Ca2+](i) due to Ca2+ influx. I-NSC plays important roles in forming membrane potentials and regulating [Ca2+](i) of CASMCs under various pathophysiological conditions such as ischemia.

KLF5/BTEB2 is a Kruppel-like zinc-finger type transcription factor of a nonmuscle type myosin heavy chain gene SMemb, which is markedly induced in phenotypically modulated smooth muscle (SMC). KLF5/BTEB2 expression in SMC is downregulated with vascular development in vivo but upregulated in neointima that is produced in response to vascular injury. KLF5/BTEB2 activates the promoter of not only SMemb gene, but also of other genes which are actvated in synthetic SMC, including plasminogen activator inhibitor-1 (PAI-1), iNOS, PDGF-A, Egr-1 and VEGF receptors. Mitogenic stimulation activates KLF5/BTEB2 gene expression through MEK1 and Egr-1. KLF5/BTEB2 +/- mice showed a marked reduction in neointimal formation, angiogenesis and cardiac fibrosis induced by various kinds of stress. We suggest that KLF5/BTEB2 is a crucial transcription factor involved in smooth muscle phenotypic modulation as well as mesenchymal cell activation in cardiovascular disease.

As the number of the genetic studies has rapidly increased in recent years, there has been growing concern that the privacy of the participants in such studies can be invaded unless effective measures are adopted to protect confidentiality. It is crucial for the scientific community to establish a method to anonymize DNA samples so that the public will trust genetic researchers. Here, we present a reliable and practical method of making DNA samples used in the genetic research anonymous. It assures complete anonymity by coding samples and personal information twice. Since it does not require equipment, such as bar-code readers or a software package, its cost is nominal compared with the laboratory costs. All institutions engaged in genetic research may wish to take measures such as the one described here to ensure the privacy and confidentiality of the participants in their genetic studies.

Background/Aims: Histamine H2 receptor antagonists are considered to exert their effects on gastric acid secretion more rapidly than proton pump antagonists. However, there are no reports concerning the direct interaction of a histamine H2 receptor antagonist with the human H2 receptor in terms of onset of action. This study aims to characterize how rapidly famotidine and ranitidine, the most widely used histamine H2 receptor antagonists, interact with the human histamine H2 receptor. Methods: HEK293 cell lines, stably expressing human histamine H2 receptors, were obtained. The dose- and time-dependent effects of famotidine and ranitidine on [3H]-tiotidine binding and histamine-stimulated cAMP production were analyzed. Results: Ranitidine inhibited both [3H]-tiotidine binding and histamine-stimulated cAMP production more promptly than did famotidine. Inhibition of histamine- stimulated cAMP production by Cmax doses of famotidine (20 mg p.o.) and ranitidine (150 mg p.o.) peaked by 15 and 2 min, respectively. [ 3H]-Tiotidine binding was not saturated by 60 min at the famotidine Cmax, while the ranitidine Cmax had produced saturation by 15 min. Conclusion: Ranitidine inhibits the human histamine H2 receptor very rapidly. Copyright © 2003 S. Karger AG, Basel.

Leptin-deficient ob/ob mice show many characteristics of obesity, including excess peripheral adiposity as well as severe hepatic steatosis, at least in part, due to increased hepatic lipogenesis. Polyunsaturated fatty acids (PUFAs) are not only ligands for peroxisome proliferator-activated receptor (PPAR) α but are also negative regulators of hepatic lipogenesis, which is thought to be mediated by the repression of sterol regulatory element-binding protein (SREBP)-1. We have previously shown that the disruption of SREBP-1 in ob/ob mice decreased their liver triglyceride storage. To examine whether PUFAs could reduce hepatic triglyceride deposition, we challenged ob/ob mice with dietary PUFA. It is demonstrated that PUFA markedly decreased the mature form of SREBP-1 protein and thereby reduced the expression of lipogenic genes such as fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1) in the livers of ob/ob mice. Consequently, the liver triglyceride content and plasma alanine aminotransferase (ALT) levels were decreased. Furthermore, both hyperglycemia and hyperinsulinemia in ob/ob mice were improved by PUFA administration, similar to the effect of PPARα activators. In conclusion, PUFAs ameliorate obesity-associated symptoms, such as hepatic steatosis and insulin resistance, presumably through both down-regulation of SREBP-1 and activation of PPARα.

We recently isolated a Krüppel-like zinc-finger transcription factor 5 (KLF5; also known as BTEB2 and IKLF), which is markedly induced in activated vascular smooth-muscle cells and fibroblasts. Here we describe our analysis of the in vivo function of KLF5 using heterozygous KLF5-knockout mice (Klf5(+/-)). In response to external stress, Klf5(+/-) mice showed diminished levels of arterial-wall thickening, angiogenesis, cardiac hypertrophy and interstitial fibrosis. Also, angiotensin II induced expression of KLF5, which in turn activated platelet-derived growth factor-A (PDGF-A) and transforming growth factor-beta (TGF-beta) expression. In addition, we determined that KLF5 interacted with the retinoic-acid receptor (RAR), that synthetic RAR ligands modulated KLF5 transcriptional activity, and that in vivo administration of RAR ligands affected stress responses in the cardiovascular system in a KLF5-dependent manner. KLF5 thus seems to be a key element linking external stress and cardiovascular remodeling.

A novel murine model of aging (kl/kl mice) has been developed by in vivo mutagenesis. We analyzed endothelial function in this strain. Ring preparations of the thoracic aorta were obtained from 6- to 9-week old wild-type (+/+) and heterozygous (kl/+) klotho mice. The aortas of kl/+ mice showed an exaggerated contractile response to norepinephrine and attenuated vasodilator responses to acetylcholine and lecithinized superoxide dismutase (SOD) compared to +/+ mice. The response to sodium nitroprusside was unaltered in kl/+ mice. The contraction in response to norepinephrine was augmented by treatment with N(G)-nitro-L-arginine methyl ester (L-NAME, 10(-5) M) to a greater extent in +/+ mice than in kl/+ mice. Treatment with L-NAME abolished the vasodilator responses to both acetylcholine and lecithinized SOD. NO metabolites (NO2- and NO3-) and cGMP concentrations in the urine were significantly reduced in kl/+ mice compared to +/+ mice. However, the urinary excretion of 6-keto-prostaglandin F1alpha was unaltered. There was little immunostaining for NO synthase and vascular endothelial growth factor (VEGF) in the aorta of kl/+ mice. No immunostaining for NO synthase was noted in the aorta of kl/kl mice. The expression of the klotho gene product may have a role in the regulation of VEGF expression and is tightly linked to endothelial release of NO.

BACKGROUND/AIMS: We previously showed that the mesangial expression of nonmuscle-type myosin heavy chain, SMemb, was related to glomerular sclerosis. Although angiotensin II (AII) is known to promote the glomerular accumulation of extracellular matrix and sclerosis, the effect of AII on the mesangial expression of SMemb is unknown. Thus, we investigated the effect of AII on the mesangial expression of SMemb and synthesis of fibronectin. METHODS: We continuously administered AII to Sprague-Dawley rats with subcutaneous osmotic minipumps for 14 days. Control animals received normal saline instead. The effects of oral administration of an AII type 1 (AT1) receptor antagonist, candesartan cilexetil, and a vasodilator, hydralazine, were also examined. RESULTS: Semiquantitative immunohistochemical evaluation and Western blot analysis showed that AII significantly enhanced glomerular expression of SMemb (0.87 +/- 0.33 vs. 0.40 +/- 0.19 for immunohistochemical grading, p < 0.05; 2.55 +/- 0.88 vs. 1.16 +/- 0.75 for Western blot analysis, p < 0.05). Glomerular fibronectin was also increased in AII-administered rats (301.7 +/- 206.8 ng/5 microg protein vs. 95.8 +/- 81.3 ng/5 microg protein, p < 0.05). Candesartan cilexetil attenuated these effects. On the other hand, hydralazine did not change the glomerular expression of SMemb enhanced by AII administration. CONCLUSION: All induced a phenotypic alteration in mesangial cells, enhanced the mesangial expression of SMemb and stimulated the fibronectin synthesis. These results suggest that the mesangial expression of SMemb is related to glomerular matrix accumulation and that AII mediates both mesangial processes via AT1 receptors.

Background/Aims: We previously showed that the mesangial expression of nonmuscle-type myosin heavy chain, SMemb, was related to glomerular sclerosis. Although angiotensin 11 (All) is known to promote the glomerular accumulation of extracellular matrix and sclerosis, the effect of All on the mesangial expression of SMemb is unknown. Thus, we investigated the effect of All on the mesangial expression of SMemb and synthesis of fibronectin. Methods: We continuously administered All to Sprague-Dawley rats with subcutaneous osmotic minipumps for 14 days. Control animals received normal saline instead. The effects of oral administration of an All type 1 (AT1) receptor antagonist, candesartan cilexetil, and a vasodilator, hydralazine, were also examined. Results: Semiquantitative immunohistochemical evaluation and Western blot analysis showed that All significantly enhanced glomerular expression of SMemb (0.87 +/- 0.33 vs. 0.40 +/- 0.19 for immunohistochemical grading, p &lt; 0.05; 2.55 +/- 0.88 vs. 1.16 +/- 0.75 for Western blot analysis, p &lt; 0.05). Glomerular fibronectin was also increased in All-administered rats (301.7 &PLUSMN; 206.8 ng/5 pg protein vs. 95.8 &PLUSMN; 81.3 ng/5 pg protein, p &lt; 0.05). Candesartan cilexetil attenuated these effects. On the other hand, hydralazine did not change the glomerular expression of SMemb enhanced by All administration. Conclusion: All induced a phenotypic alteration in mesangial cells, enhanced the mesangial expression of SMemb and stimulated the fibronectin synthesis. These results suggest that the mesangial expression of SMemb is related to glomerular matrix accumulation and that All mediates both mesangial processes via AT1 receptors. Copyright (C) 2002 S. Karger AG, Basel.

Mechanical stress activates various hypertrophic responses, including activation of mitogen-activated protein kinases (MAPKs) in cardiac myocytes. Stretch activated extracellular signal-regulated kinases partly through secreted humoral growth factors, including angiotensin II, whereas stretch-induced activation of c-Jun NH(2)-terminal kinases and p38 MAPK was independent of angiotensin II. In this study, we examined the role of integrin signaling in stretch-induced activation of p38 MAPK in cardiomyocytes of neonatal rats. Overexpression of the tumor suppressor PTEN, which inhibits outside-in integrin signaling, strongly suppressed stretch-induced activation of p38 MAPK. Overexpression of focal adhesion kinase (FAK) antagonized the effects of PTEN, and both tyrosine residues at 397 and 925 of FAK were necessary for its effects. Stretch induced tyrosine phosphorylation and activation of FAK and Src. Stretch-induced activation of p38 MAPK was abolished by overexpression of FAT and CSK, which are inhibitors of the FAK and Src families, respectively, and was suppressed by overexpression of a dominant-negative mutant of Ras. Mechanical stretch-induced increase in protein synthesis was suppressed by SB202190, a p38 MAPK inhibitor. These results suggest that mechanical stress activates p38 MAPK and induces cardiac hypertrophy through the integrin-FAK-Src-Ras pathway in cardiac myocytes.

Na/Ca交換体(NCX)は,心筋細胞内からのCa2+汲み出しに中心的役割を果たす膜上タンパクであるが,その機能異常がどのような影響を生体に及ぼすのかは判っていない.今回我々は,NCXノックアウトヘテロ接合体(NCX KO)マウスと正常な同胞(WT)を用いてNCX機能異常に関する検討を行った.単離心室筋細胞を電気刺激し,細胞内Ca2+濃度([Ca2+]i)変化を観察した(fluo-3,室温).誘発された[Ca2+]iトランジェントの振幅はNCX KOで有意に大きく,続いてcaffeine(20mM)処理をして測定した筋小胞体(SR)内Ca2+量は,NCX KOで著明に増加していた.ノーザンブロットによるL型Ca2+チャネルの発現レベルに差がないことから,電気刺激時の[Ca+]i振幅の上昇はSR内Ca2+量の増加によるCa2+依存性Ca+放出の増強によるためと考えられた.これらの結果から,NCX KOマウス心室筋細胞はCa2+-overloadに陥りやすいと考えられた.横行大動脈部分に縮窄を形成し心肥大を誘発したところ,NCX KO心においてより顕著な肥大が観察された.心内圧測定の結果から,心内圧上昇がNCX KOマウスにおいてより大きかったことがその原因と推察された.血行動態上の拡張期dip&plateauパターンやドップラーエコー上の左室拡張早期流入血流減速時間(DCT)の短縮といった拡張不全所見を認めるなど,NCX KOにて心肥大が著明であることを示唆する生理学的所見も得られた.一方, WTではすでに収縮不全(EF低下)を呈しSR CaATPase(SERCA)の発現の低下も伴っていることから,心不全へ移行しているものと思われた.総括すると,NCX機能の低下は,圧負荷に対する肥大反応は加速するものの心不全への移行に関しては抑制的に作用すると考えられる.不全心ではNCXの発現および機能が上昇しているとされているが,このことからも心不全の進行にNCX機能変化が大きく関与する可能性が示唆された.

The aim of this study was to test the hypothesis that a subendocardial arrhythmogenic focus makes the heart more susceptible to VF due to electrical interaction with the Purkinje network. Monofocal ventricular tachycardia (mVT) was created by injecting 5-μg aconitine into the left ventricular subepicardium (EPI-mVT, n = 8) or subendocardium (ENDO-mVT, n = 13) in anesthetized dogs. Despite the similar cycle length of mVT, the incidence of VF was significantly different between EPI-mVT and ENDO-mVT (100 [8/8] vs 46% [6/13], P &lt 0.05). VF was invariably preceded by hemodynamic deterioration. Three-dimensional cardiac mapping (n = 10, 221 ± 11 recording sites) revealed that VF was triggered solely by focal firing unrelated to the primary arrhythmogenic focus in both mVT models. No interaction between the primary focus and adjacent endocardial tissue was indicated. These results suggest that the proximity of the arrhythmogenic focus to the Purkinje network has little role in cardiac vulnerability to VF, and that progression of mVT to VF is largely caused by sporadic focal firing regardless of the site of the arrhythmogenic focus in the present animal model.

Accelerated coronary arteriosclerosis remains a major problem for the long-term survival of cardiac transplant recipients. However, the pathogenesis of graft vasculopathy is poorly understood and there is no effective therapy. Tranilast is a promising drug that may prevent postangioplasty restenosis. Here, we investigated whether orally administered tranilast inhibits the development of intima hyperplasia in a mouse model of cardiac transplantation. Cardiac allografts from BALB/c mice were transplanted heterotopically into C3H/He mice. Mice were administered either vehicle or tranilast everyday by gavage. Morphometrical analysis of the cardiac allografts harvested at 2 months revealed that the administration of tranilast significantly reduced the development of coronary atherosclerosis. In the mice treated with tranilast, upregulation of the cyclin-dependent kinase inhibitor p21 was observed in the allografts, accompanied by a reduced number of proliferating cells. Tranilast also suppressed transforming growth factor-beta (TGF-beta) expression. Tranilast may be effective in preventing transplant-associated arteriosclerosis through its anti-inflammatory and anti-proliferative effects. (C) 2001 Elsevier Science B.V All rights reserved.

BACKGROUND: We recently suggested that a rat aortotomy model could be substituted for a vascular anastomotic stricture around a suture line. The aim of this study was to verify such a rat aortotomy model using proliferating cell nuclear antigen (PCNA) immunohistochemistry, which is a sensitive method for detection of proliferating cells in vascular tissue after injury. METHODS: Longitudinal aortotomy was performed in the abdominal aorta of the rat subjects. The rats were sacrificed at 1, 2, 4 and 8 weeks following aortotomy (n=20 in each group). The control rats were sacrificed without aortotomy (n=20). The percentage of the lumen occluded by intimal thickening (I/M ratio) was calculated. All tissues were stained with antibodies against proliferating cell nuclear antigen (PCNA). RESULTS: Values of I/M ratio were 8.5+/-4.4%, 15.6+/-9.5%, 11.8+/-5.5%, 9.6+/-6.2% at 1, 2, 4, 8 weeks following aortotomy in the injured groups and 0.4+/-0.1% in the controls, respectively. Those values in the injured group increased significantly as compared to the controls. There were also significant differences between one week and two weeks following aortotomy. The PCNA labeling index at one week following aortotomy (21.4+/-2.7%) was significantly higher than at two weeks following aortotomy (2.8+/-1.9%). SMCs in the intima at four and eight weeks following aortotomy were completely negative for PCNA. CONCLUSIONS: The experimental rat aortotomy model was determined to be useful in the investigation of intimal thickening around the suture line.

Peroxisome proliferator-activated receptor (PPAR) gamma is a ligand-activated transcription factor and a member of the nuclear hormone receptor superfamily that is thought to be the master regulator of fat storage; however, the relationship between PPARgamma and insulin sensitivity is highly controversial. We show here that supraphysiological activation of PPARgamma by PPARgamma agonist thiazolidinediones (TZD) markedly increases triglyceride (TG) content of white adipose tissue (WAT), thereby decreasing TG content of liver and muscle, leading to amelioration of insulin resistance at the expense of obesity. Moderate reduction of PPARgamma activity by heterozygous PPARgamma deficiency decreases TG content of WAT, skeletal muscle, and liver due to increased leptin expression and increase in fatty acid combustion and decrease in lipogenesis, thereby ameliorating high fat diet-induced obesity and insulin resistance. Moreover, although heterozygous PPARgamma deficiency and TZD have opposite effects on total WAT mass, heterozygous PPARgamma deficiency decreases lipogenesis in WAT, whereas TZD stimulate adipocyte differentiation and apoptosis, thereby both preventing adipocyte hypertrophy, which is associated with alleviation of insulin resistance presumably due to decreases in free fatty acids, and tumor necrosis factor alpha, and up-regulation of adiponectin, at least in part. We conclude that, although by different mechanisms, both heterozygous PPARgamma deficiency and PPARgamma agonist improve insulin resistance, which is associated with decreased TG content of muscle/liver and prevention of adipocyte hypertrophy.

To elucidate pathophysiological roles of heme oxygenase (HO)-1 in regulation of vascular tone in vivo, we have developed and characterized transgenic (Tg) mice that overexpress HO-1 site specifically in vascular smooth muscle cells (VSMCs). The Tg mice were generated by use of human HO-1 cDNA under the control of SM22-alpha promoter. The HO-1 gene overexpression was demonstrated by Northern blot analysis and coincided with increases in the protein expression in VSMCs and total HO activities. Tg mice exhibited a significant increase in arterial pressure at various ages and displayed impaired nitrovasodilatory responses in isolated aortic segments versus nontransgenic littermates while enhancing their nitric oxide (NO) production. The pressure of Tg mice was unchanged by systemic administration of either N(omega)-nitro-L-arginine or SNP. Furthermore, the isolated aorta in these mice exhibited lesser extents of NO-elicited cGMP elevation via soluble guanylate cyclase (sGC), while exhibiting no notable downregulation of sGC expression. Such impairment of the NO-elicited cGMP increase was restored significantly by tin protoporphyrin IX, an HO inhibitor. On the other hand, 3-(5'-hydroxymethyl-2' furyl)-1-benzyl-indazol (YC-1), an NO-independent activator of sGC, increased cGMP and relaxed aortas from Tg mice to levels comparable with those from nontransgenic mice, which indicates that contents of functionally intact sGC are unlikely to differ between the two systems. These findings suggest that site-specific overexpression of HO-1 in VSMCs suppresses vasodilatory response to NO and thereby leads to an elevation of arterial pressure.

Recently, troglitazone has emerged as an insulin sensitizer for the treatment of type II diabetes. However, its effect on skeletal muscle glucose use (SMGU) has not been studied. Methods: To investigate the effect of troglitazone on SMGU in patients with type II diabetes, we undertook skeletal muscle F-18-FDG PET dynamic imaging under insulin clamping before and after administration of SMGU to 20 patients with type Ii diabetes. Data were compared with those for 12 age-matched healthy volunteers. Results: The whole-body glucose disposal rate (GDR) was significantly lower in patients 129.9 +/- 9.83 mu mol/min/kg) than in control subjects (55.6 +/- 16.5 mu mol/min/kg, P &lt; 0.01), as was the SMGU (patients, 3.27 +/-: 2.17 mu mol/min/kg; control subjects, 10.9 +/- 6.4 mu mol/min/kg; P &lt; 0.01). After the therapy, GDR significantly improved in patients (29.3 +/- 14.6 mu mol/min/kg, P &lt; 0.05), as did SMGU (5.96 +/- 2.11 mu mol/min/kg, P &lt; 0.05). When results for patients with and without hypertension were separately analyzed, a significant improvement in SMGU after troglitazone was seen in both normotensive and hypertensive patients 9 normotensive [n = 10]: baseline, 3.67 +/- 2.89 mu mol/min/kg; after therapy, 5.28 +/- 2.57 mu mol/min/kg; P &lt; 0.05; hypertensive [n = 10]: baseline, 2.89 +/- 1.22 mu mol/min/kg; after therapy, 4.72 +/- 1.39 mu mol/min/kg; P &lt; 0.05). GDR in patients with and without hypertension was significantly improved by troglitazone (normotensive: baseline, 17.9 +/- 10.2 mu mol/min/kg; after therapy, 31.9 +/- 15.9 mu mol/min/kg; P &lt; 0.01; hypertensive: baseline, 39.6 +/- 15.1 mu mol/min/kg; after therapy, 47.7 +/- 23.8 mu mol/min/kg; P &lt; 0.05). The plasma free fatty acid concentration during insulin clamping was not changed by troglitazone (baseline, 1.1 +/- 0.86 mEq/L; after therapy, 0.93 +/- 0.65 mEq/L; P = not significant). Conclusion: Troglitazone can improve whole-body insulin resistance through the improvement of SMGU but not through a decline in plasma free fatty acid concentration in patients with type II diabetes with or without hypertension.

gp130, a common receptor for the interleukin 6 family, plays pivotal roles in growth and survival of cardiac myocytes. In the present study, we examined the role of gp130 in pressure overload-induced cardiac hypertrophy using transgenic (TG) mice, which express a dominant negative mutant of gp130 in the heart under the control of alpha myosin heavy chain promoter. TG mice were apparently healthy and fertile. There were no differences in body weight and heart weight between TG mice and littermate wild type (WT) mice. Pressure overload-induced increases in the heart weight/body weight ratio, ventricular wall thickness, and cross-sectional areas of cardiac myocytes were significantly smaller in TG mice than in WT mice. Northern blot analysis revealed that pressure overload-induced up-regulation of brain natriuretic factor gene and down-regulation of sarcoplasmic reticulum Ca(2+) ATPase 2 gene were attenuated in TG mice. Pressure overload activated ERKs and STAT3 in the heart of WT mice, whereas pressure overload-induced activation of STAT3, but not of ERKs, was suppressed in TG mice. These results suggest that gp130 plays a critical role in pressure overload-induced cardiac hypertrophy possibly through the STAT3 pathway.

To determine the presence and functional role of the histamine H2 receptor (H2R) palmitoylation, a receptor with a Cys(305) to Ala (A(305) receptor) mutation was generated. Wild-type (WT) and A(305) receptors were tagged at their N-termini with a hemagglutinin (HA) epitope. WT, but not A(305). receptors incorporated [H-3]palmitate by metabolic labeling, indicating that the H2R is palmitoylated at Cys(305). Immunocytochemistry of WT and A(305) receptors expressed in COS7 cells revealed WT receptors to be distributed at the plasma membrane, while the majority of A(305) receptors were localized intracellularly with only a small portion being at the plasma membrane. However, the affinity of the A(305) receptor for tiotidine was comparable to that of the WT receptor. In addition. when the amounts of cell surface receptors as determined by anti-HA antibody binding were equivalent, A(305) receptors mediated production of more cAMP than WT receptors. Preincubation of COS7 cells expressing each receptor with 10(-5) M histamine for 30 min reduced subsequent cAMP production in response to histamine via the receptors to similar extents, indicating that palmitoylation is not necessary for desensitization. In addition, cell surface A(305) receptors were capable of being internalized from the cell surface at a rate and extent similar to those of WT receptors. Finally, CHO cell lines stably expressing either WT or A(305) receptors were incubated with 10(-5) M histamine for 1, 6, 12 and 24 h. Total amounts of WT and A305 receptors, as determined by tiotidine binding, were reduced by incubation, indicating downregulation. Downregulation of the A(305) receptor was more extensive than that of the WT receptor. Thus, palmitoylation of the H2R might be important for targeting to the cell surface and stability. (C) 2001 Elsevier Science B.V. All rights reserved.

BACKGROUND: It remains unclear how hemodynamic overload induces cardiac hypertrophy. Recently, activation of calcium-dependent phosphatase, calcineurin, has been elucidated to induce cardiac hypertrophy. In the present study, we examined the role of calcineurin in load-induced cardiac hypertrophy by using Dahl salt-sensitive (DS) rats, which develop both pressure and volume overload when fed a high salt diet. METHODS AND RESULTS: In the DS rat heart, the activity of calcineurin was increased and cardiac hypertrophy was induced by high salt diet. Treatment of DS rats with the calcineurin inhibitor FK506 (0.1 or 0.01 mg/kg every second day) from the age of 6 weeks to 12 weeks inhibited the activation of calcineurin in the heart in a dose-dependent manner and attenuated the development of load-induced cardiac hypertrophy and fibrosis without change of hemodynamic parameters. Additionally, treatment with 0.1 mg/kg every second day but not with 0.01 mg/kg every second day of FK506 from the age of 12 weeks to 16 weeks induced regression of cardiac hypertrophy in DS rats. Load-induced reprogramming of gene expression was also suppressed by the FK506 treatment. CONCLUSIONS: These results suggest that calcineurin is involved in the development of cardiac hypertrophy in rats with salt-sensitive hypertension and that inhibition of calcineurin could induce regression of cardiac hypertrophy.

We studied whether plasma BNP concentration measurement was useful in screening for silent ischemia or asymptomatic cardiac dysfunctions in diabetic outpatients in daily clinical practice. Plasma BNP and ANP concentrations in 573 outpatients with diabetes were measured. Plasma BNP concentrations increased with complications of hypertension, heart diseases, cardiac dysfunction, diabetic neuropathy, diabetic nephopathy, and diabetic retinopathy. Plasma BNP also was significantly correlated with NYHA classification, blood pressure, cardio-thoracic-ratio (CTR), SV1 + SV5, 6, interventricular septum thickness, posterior wall thickness, left ventricular mass, and left ventricular mass index. In subjects without heart disease, plasma ANP concentrations showed a correlation with blood glucose, but plasma BNP concentrations did not show such a correlation. Receiver operating characteristic analysis revealed that plasma BNP concentrations showed sensitivity of 62.7% and specificity of 75.0% in detecting heart diseases or cardiac dysfunction. In addition, the result of logistic regression analysis indicated that plasma BNP measurement was the best clinical parameter in the prediction of heart diseases and cardiac dysfunction. Measurement of the plasma BNP concentrations is thus useful in diagnosing heart diseases in diabetic subjects in daily clinical practice. © 2001, THE JAPAN DIABETES SOCIETY. All rights reserved.

We report a case of intravascular lymphoma (IVL) complicated with Pneumocystis carinii pneumonia (PCP). A 65-year-old male complaining of dyspnea and dementia was diagnosed to have pulmonary IVL by transbronchial lung biopsy. Concomitantly, deoxyribonucleic acid sequence specific to Pneumocystis carinii was detected in bronchoalveolar lavage fluid by polymerase chain reaction. Differential responses to the sequential treatments for PCP and IVL implied that increased serum lactate dehydrogenase (LDH) was due to PCP, whereas hypoxemia and dementia were due to IVL. Although pulmonary IVL and PCP share many clinical presentations, exact diagnosis is essential for their successful treatment.

Previous studies with peritoneal macrophages obtained from macrophage scavenger receptor class A (MSR-A) knock-out mice showed that the endocytic uptake of advanced glycation end products (AGE) by macrophages was mediated mainly by MSR-A. However, it is controversial whether the endocytic uptake of intravenously injected AGE proteins by liver sinusoidal endothelial cells (LECs) is similarly explained by receptor-mediated endocytosis via MSR-A, The present study was conducted to compare the capacity to endocytose AGE proteins in LECs and peritoneal macrophages obtained from MSR-A knockout and littermate wild-type mice. The endocytic degradation capacity of MSR-A knock-out LECs for AGE-BSA was indistinguishable from that of wild-type LECs, whereas that of MSR-A knock-out peritoneal macrophages for AGE-BSA was decreased to 30% of that in wild-type cells. Similarly, the endocytic degradation of MSR-A knock-out LECs for acetylated low-density lipoprotein (acetyl-LDL) did not differ from that of wild-type LECs, whereas the endocytic degradation of acetyl-LDL by MSR-A knock-out peritoneal macrophages was less than 20% of that in wild-type cells. Furthermore, formalde-hydetreated serum albumin (f-Alb), a ligand known to undergo scavenger-receptor-mediated endocytosis by LECs, was effectively taken up by MSR-A knock-out LECs at a capacity that did not differ from that of wild-type LECs. Moreover, the endocytic uptake of AGE-BSA by LECs was effectively competed for by unlabelled f-Alb or acetyl-LDL. These results indicate that the scavenger-receptor ligands AGE proteins, acetyl-LDL and f-Alb are endocytosed by LECs through a non-MSR-A pathway.