Masaoki Takano

Publications of Masaoki Takano

• Phosphoproteome profiling using a fluorescent phosphosensor dye in two-dimensional polyacrylamide gel electrophoresis.

  Authors: Mieko Otani, Taizo Taniguchi, Akiko Sakai, Jouji Seta, Keiichi Kadoyama, Tooru Nakamura-Hirota, Shogo Matsuyama, Keiji Sano, Masaoki Takano

  Applied biochemistry and biotechnology. 03/2011; 164(6):804-18.

  We validated the novel PhosphoQUANTI SolidBlue Complex (PQSC) dye for the sensitive fluorescent detection of phosphorylated proteins in polyacrylamide- and two-dimensional gel electrophoresis (PAGEWe validated the novel PhosphoQUANTI SolidBlue Complex (PQSC) dye for the sensitive fluorescent detection of phosphorylated proteins in polyacrylamide- and two-dimensional gel electrophoresis (PAGE and 2DE, respectively). PQSC can detect as little as 15.6 ng of ß-casein, a pentaphosphorylated protein, and 61.3 ng of ovalbumin, a diphosphorylated protein. Fluorescence intensity correlates with the number of phosphorylated residues on the protein. To demonstrate the specificity of PQSC for phosphoproteins, enzymatically dephosphorylated lysates of Swiss 3T3 cells were separated in 2DE gels and stained by PQSC. The fluorescence signals in these gels were markedly reduced following dephosphorylation. When the phosphorylated proteins in Swiss 3T3 cell lysates were concentrated using a phosphoprotein enrichment column, the majority of phosphoproteins showed fluorescence signals in the pI 4-5 range. Finally, we performed phosphoproteome analysis to study differences in the protein phosphorylation profiles of proliferating and quiescent Swiss 3T3 cells. Over 135 discernible protein spots were detected, from which a selection of 15 spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF-MS). The PQSC staining procedure for phosphoprotein detection is simple, reversible, and fully compatible with MALDI TOF-MS.

• Synaptotagmin1 synthesis induced by synaptic plasticity in mouse hippocampus through activation of nicotinic acetylcholine receptors.

  Authors: Takaaki Nishimoto, Keiichi Kadoyama, Taizo Taniguchi, Masaoki Takano, Mieko Otani, Tooru Nakamura-Hirota, Yabin Lu, Akira Matsumoto, Shogo Matsuyama

  Neuroscience letters. 02/2011; 489(1):25-9.

  We have reported that systemic application of nicotinic agonists expresses a long-term potentiation (LTP)-like facilitation, a model of synaptic plasticity, in vivo in the mouse hippocampus. TheWe have reported that systemic application of nicotinic agonists expresses a long-term potentiation (LTP)-like facilitation, a model of synaptic plasticity, in vivo in the mouse hippocampus. The present study conducted to clarify the involvement of synaptotagmin1 in synaptic plasticity by investigating the time-dependent change of the mRNA and protein levels of synaptotagmin1 during LTP-like facilitation in the mouse hippocampus. The mRNA expression of synaptotagmin1 increased during 2- to 8-h period by intraperitoneal application of nicotine (3mg/kg), returning to the basal level in 12-h. Also, the protein level of synaptotagmin1, but not synaptophysin, in a total fraction from hippocampus increased during 4- to 12-h period by the same treatment, returning to the basal level in 24-h. The protein level of synaptotagmin1 in a membrane fraction from hippocampus also increased during 4- to 8-h period by nicotine, returning to the basal level in 12-h. This nicotine-enhanced synaptotagmin1 protein in a membrane fraction was inhibited by pretreatment of mecamylamine (0.3mg/kg, i.p.), a nonselective nicotinic acetylcholine receptors (nAChRs) antagonist. Furthermore, choline (30mg/kg, i.p.), a selective α7 nAChR agonist, or ABT-418 (10mg/kg, i.p.), a selective α4β2 nAChR agonist, enhanced the level of synaptotagmin1 in a membrane fraction. Our findings demonstrate that synaptotagmin1 protein following mRNA which is enhanced without increasing the number of synapse gathers around pre-synaptic membrane during hippocampal LTP-like facilitation through activation of α7 and/or α4β2 nAChRs in the brain. These results suggest that new-synthesized synaptotagmin1 following synaptic plasticity may contribute to long-lasting synaptic plasticity via positive, feedfoward mechanisms.

• β-Citryl-L-glutamate acts as an iron carrier to activate aconitase activity.

  Authors: Michiko Hamada-Kanazawa, Masanori Narahara, Masaoki Takano, Kyong Son Min, Keiichi Tanaka, Masaharu Miyake

  Biological & pharmaceutical bulletin. 01/2011; 34(9):1455-64.

  The compound β-citryl-L-glutamate (β-CG) was initially isolated from developing brains, though its functional roles remain unclear. In in vitro experiments, the [Fe(II)(β-CG)] complex activatedThe compound β-citryl-L-glutamate (β-CG) was initially isolated from developing brains, though its functional roles remain unclear. In in vitro experiments, the [Fe(II)(β-CG)] complex activated aconitase in the presence of reducing reagents, whereas no Fe complex with citrate, glutamate, or deferoxamine displayed such an effect. β-CG and [Fe(II)(β-CG)] both bound to the fourth labile Fe atom (Fe(a)) in the [4Fe-4S] cluster of aconitase. Furthermore, [Fe(II)(β-CG)] reactivated aconitase damaged by ammonium peroxodisulfate (APS), while β-CG and citrate had no effect. These findings suggest that [Fe(II)(β-CG)] can transfer Fe to aconitase disassembled by APS. In intact mitochondria, both β-CG and [Fe(II)(β-CG)] bound to Fe(a) of aconitase, whereas only [Fe(II)(β-CG)] reactivated the enzyme disassembled by APS. In cultured neuronal cells, β-CG significantly enhanced cell viability by accelerating mitochondrial activity in primary cultures of neurons from newborn mouse cerebrum tissues. Thus, the β-CG plays a role as an Fe-carrier for mitochondrial aconitase, and then activates it. Taken together, these findings suggest that β-CG is an endogenous low molecular weight Fe chaperone for aconitase.

• Use of a phosphosensor dye in proteomic analysis of human mutant tau transgenic mice.

  Authors: Masaoki Takano, Mieko Otani, Akiko Sakai, Keiichi Kadoyama, Shogo Matsuyama, Akira Matsumoto, Mariko Takenokuchi, Miho Sumida, Taizo Taniguchi

  Neuroreport. 11/2009;

  Recently, we have generated transgenic mice (designated as SJLB) carrying human N279K mutant tau, one of the tau mutations causing parkinsonism linked to chromosome 17 (FTDP-17). SJLB mice mimic someRecently, we have generated transgenic mice (designated as SJLB) carrying human N279K mutant tau, one of the tau mutations causing parkinsonism linked to chromosome 17 (FTDP-17). SJLB mice mimic some features of behavioral alterations and neuronal pathology of patients with Alzheimer's disease. To investigate how tau dysfunctions cause these features, we examined the expression and phosphorylation levels in SJLB mouse hippocampal proteins using a phosphosensor dye in two-dimensional poly acrylamide gel electrophoresis analysis and mass spectrometry. Calreticulin and tubulin beta4 are significantly more phosphorylated, and heat shock cognate 71 kDa protein, tubulin beta2, vacuolar ATP synthase catalytic subunit A, alpha-internexin, alpha-enolase, ubiquitin carboxyl-terminal hydrolase isozyme L1, and complexin-2 are significantly less phosphorylated in SJLB mice than control mice. These proteins could be new targets for elucidating underlying mechanisms and therapeutic intervention in neurodegenerative diseases.

• Germ cell specific protein VASA is over-expressed in epithelial ovarian cancer and disrupts DNA damage-induced G2 checkpoint.

  Authors: Hisashi Hashimoto, Tamotsu Sudo, Yoshiki Mikami, Mieko Otani, Masaoki Takano, Hiroshi Tsuda, Hiroaki Itamochi, Hidetaka Katabuchi, Masaharu Ito, Ryuichiro Nishimura

  Gynecologic oncology. 10/2008;

  OBJECTIVE: Cancer cells have characteristics, such as high telomerase activity and high levels of migration activity and proliferation, which are very similar to those of germ cell lineages. In thisOBJECTIVE: Cancer cells have characteristics, such as high telomerase activity and high levels of migration activity and proliferation, which are very similar to those of germ cell lineages. In this study, we examined the expression of VASA, a germ cell lineage specific marker and evaluated its clinical significance in epithelial ovarian cancer (EOC). METHODS: We investigated VASA expression in 75 EOC tissues by immunohistochemistry, correlating results with clinicopathological factors. To clarify the effects of VASA on cellular phenotypes, we compared the protein expression profiles between SKOV-3 cells stably expressing VASA (SKOV-3-VASA) and vector-control cell lines by coupling 2D fingerprinting and identification of proteins by mass spectrometry. RESULTS: VASA expression in tumor cells was found in 21of 75 cases and was positively correlated with high age and serous histology. Significant down-regulation of 14-3-3sigma was observed in SKOV-3-VASA versus control cells. Over-expression of VASA abrogates the G2 checkpoint, induced by DNA damage, by down-regulating the expression of 14-3-3sigma. CONCLUSIONS: These results suggest that VASA may either play a direct role in the progression of EOC or serve as a valuable marker of tumorigenesis.

• Lipopolysaccharide activates the kallikrein-kinin system in mouse choroid plexus cell line ECPC4.

  Authors: Masaoki Takano, Chieko Satoh, Naomi Kunimatsu, Mieko Otani, Michiko Hamada-Kanazawa, Masaharu Miyake, Kyongsong Ming, Katsutoshi Yayama, Hiroshi Okamoto

  Neuroscience letters. 05/2008; 434(3):310-4.

  Regulation of the kallikrein-kinin system in cerebral inflammation is still unclear. Here, we used reverse-transcription polymerase chain reaction (RT-PCR) techniques to show that lipopolysaccharideRegulation of the kallikrein-kinin system in cerebral inflammation is still unclear. Here, we used reverse-transcription polymerase chain reaction (RT-PCR) techniques to show that lipopolysaccharide (LPS) activates the kallikrein-kinin system by enhancing liberation of bradykinin (BK), and alters mRNA levels of kallikrein-kinin system components, including high molecular weight (H-) and low molecular weight (L-) kininogens, in ECPC4 cells, a cell line of mouse choroid plexus epithelium. LPS treatment increased liberation of immunoreactive bradykinin in the supernatant of ECPC4 cells, and addition of LPS (500 ng/ml) to cultures resulted in elevation of H- and L-kininogen mRNA levels in ECPC4 cells within 24-48 h. Furthermore, LPS treatment elevated bradykinin type 2 and type 1 receptor mRNA levels within 4h, but did not change tissue kallikrein or plasma kallikrein mRNA levels. On the other hand, expression of pro-inflammatory mediators interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), and cyclooxygenase-2 mRNA increased within 4-8h after addition of LPS to ECPC4 cells. The addition of IL-1beta and TNF-alpha to investigate the major mediator for kininogen expression in ECPC4 cells remarkably induced expression of H- and L-kininogen mRNAs in ECPC4 cells. These results suggest that LPS activates the kallikrein-kinin system in the choroid plexus via autocrine induction of IL-1beta and TNF-alpha.

• Angiotensin type 2 receptor-mediated phosphorylation of eNOS in the aortas of mice with 2-kidney, 1-clip hypertension.

  Authors: Hiromi Hiyoshi, Katsutoshi Yayama, Masaoki Takano, Hiroshi Okamoto

  Hypertension. 06/2005; 45(5):967-73.

  To evaluate the role of vascular angiotensin II (Ang II) type 2 (AT2) receptor in renovascular hypertension, we investigated expressions of AT2 receptor and endothelial nitric oxide synthase (eNOS)To evaluate the role of vascular angiotensin II (Ang II) type 2 (AT2) receptor in renovascular hypertension, we investigated expressions of AT2 receptor and endothelial nitric oxide synthase (eNOS) in thoracic aortas of mice with 2-kidney, 1-clip (2K1C) hypertension. The mRNA levels of AT2 receptor in aortas, but not those of AT1 and bradykinin B2 receptors, increased 14 days but not 42 days after clipping. The contractile response to Ang II (>0.1 micromol/L) was attenuated in aortic rings excised 14 days after clipping and was restored to that of rings from sham mice by antagonists of AT2 receptor (PD123319) and B2 receptor (icatibant). The aortic levels of total eNOS, phosphorylated eNOS at Ser1177 (p-eNOS), total Akt, and phosphorylated Akt at Ser473 (p-Akt) were increased in 2K1C mice on day 14, whereas only eNOS levels were increased on day 42. The aortic cGMP levels were 20-fold greater in 2K1C mice on day 14 compared with sham mice. Administration of nicardipine for 4 days before the excision of aortas 14 days after clipping not only reduced blood pressure but also decreased the aortic levels of eNOS, p-eNOS, Akt, p-Akt, and cGMP to sham levels, whereas the administration of PD123319 or icatibant to 2K1C mice decreased p-eNOS and cGMP to sham levels without affecting blood pressure and the levels of eNOS, Akt and p-Akt. These results suggest that vascular NO production is enhanced by increased eNOS phosphorylation via the activation of AT2 receptors in the course of 2K1C hypertension.

• [Structure and signaling pathways of kinin receptors]

  Authors: Hiroshi Okamoto, Masaoki Takano

  Nippon rinsho. Japanese journal of clinical medicine. 04/2005; 63 Suppl 3:369-74.

• Stimulation of cyclic GMP production via AT2 and B2 receptors in the pressure-overloaded aorta after banding.

  Authors: Hiromi Hiyoshi, Katsutoshi Yayama, Masaoki Takano, Hiroshi Okamoto

  Hypertension. 07/2004; 43(6):1258-63.

  Abdominal aortic banding induces upregulation of the angiotensin II (Ang II) type-2 (AT2) receptor, thereby decreasing the contractile response to Ang II in the thoracic aorta of the rat. The aim ofAbdominal aortic banding induces upregulation of the angiotensin II (Ang II) type-2 (AT2) receptor, thereby decreasing the contractile response to Ang II in the thoracic aorta of the rat. The aim of this study was to use a mouse model to clarify the mechanisms by which the banding elicits upregulation of the aortic AT2 receptor and the subsequent attenuation of Ang II responsiveness. Concomitantly with the elevation in blood pressure and plasma renin concentration after banding, AT2-receptor mRNA levels in the thoracic aorta rapidly increased in mice within 4 days. Upregulation of the AT2 receptor, as well as blood pressure elevation after banding, was abolished by losartan administration. The contractile response to Ang II was depressed in aortic rings of banding mice but not of sham mice, and was restored by either the AT2-receptor antagonist PD123319 or the bradykinin B2-receptor antagonist icatibant. cGMP content in the thoracic aorta of banding mice was 9-fold greater than that of sham mice, and the elevation was reduced to sham levels 1 hour after intravenous injection of PD123319 or icatibant. When aortic rings were incubated with Ang II, cGMP content increased in banding rings but not in sham rings; the pretreatment with PD123319 or icatibant inhibited Ang II-induced cGMP production. These results suggest that aortic banding induces upregulation of the AT2 receptor through increased circulating Ang II via the AT1 receptor, thereby activating a vasodilatory pathway in vessels through the AT2 receptor via the kinin/cGMP system.

• Up-regulation of angiotensin II type 2 receptor in rat thoracic aorta by pressure-overload.

  Authors: Katsutoshi Yayama, Miyuki Horii, Hiromi Hiyoshi, Masaoki Takano, Hiroshi Okamoto, Satomi Kagota, Masaru Kunitomo

  The Journal of pharmacology and experimental therapeutics. 03/2004; 308(2):736-43.

  We have examined whether expression of angiotensin II (Ang II) type 1 (AT(1)) and/or type 2 (AT(2)) receptors are changed in thoracic aorta under pressure-overload by abdominal aortic banding in ratsWe have examined whether expression of angiotensin II (Ang II) type 1 (AT(1)) and/or type 2 (AT(2)) receptors are changed in thoracic aorta under pressure-overload by abdominal aortic banding in rats and determined whether their changes are accompanied by alteration in contractile response of thoracic aorta to Ang II. AT(2) receptor mRNA levels determined by reverse transcription-polymerase chain reaction or quantitative real-time polymerase chain reaction were increased by about 300% in aortas 4, 7, 14, and 28 days after banding without changes in AT(1) receptor mRNA levels. Contractile response of aortic rings to Ang II was decreased in thoracic aortas 7 days after banding, and AT(2) receptor antagonist PD123319 (1-[[4-(dimethulamino)-3-methylphenyl]methyl]-5-(diphenylacetyl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine-6-carboxylic acid ditrifluoroacetate) (10(-6) M) increased the Ang II responsiveness in pressure-loaded but not in sham rings. After removal of the endothelium or treatment with N(G)-nitro-L-arginine methyl ester (L-NAME), no differences were observed in Ang II responsiveness between sham and pressure-loaded rings. Either losartan (1 mg/kg/day i.p.) or candesartan (2 mg/kg/day p.o.) for 7 days after banding not only abolished the up-regulation of AT(2) receptor mRNA in aortas but also recovered their Ang II responsiveness. Basal cGMP levels were 2 times higher in pressure-loaded than in sham rings; both levels were not affected by Ang II (10(-7) M; 5 min), but greatly decreased by L-NAME (10(-4) M, 30 min). These results suggest that pressure-overload induces the up-regulation of AT(2) receptor expression in aortas via AT(1) receptor and thereby negatively modulates the vasoconstrictor sensitivity to Ang II, probably mediated by the mechanisms independent of the nitric oxide-cGMP system.

• Lipopolysaccharide injection into the cerebral ventricle evokes kininogen induction in the rat brain.

  Authors: Masaoki Takano, Masato Horie, Katsutoshi Yayama, Hiroshi Okamoto

  Brain research. 08/2003; 978(1-2):72-82.

  Kinins, such as bradykinin and Lys-bradykinin, are important mediators in peripheral inflammation. Although the existence of the components necessary for generating kinins has been demonstrated inKinins, such as bradykinin and Lys-bradykinin, are important mediators in peripheral inflammation. Although the existence of the components necessary for generating kinins has been demonstrated in the brain, a functional role of the kinin-generating system in cerebral inflammation remains to be defined. The aim of the present study was to elucidate whether inflammatory stimuli alter the mRNA levels of components for the kallikrein-kinin system, including kallikreins, kininogens and bradykinin type 2 (B(2)-) receptor in rat brain using the reverse transcription polymerase chain reaction. The intracerebroventricular (i.c.v.) injection of lipopolysaccharide (LPS; 0.25 microg/animal) resulted in the elevation of T-kininogen and high-molecular-weight (H-) kininogen mRNAs in various brain regions within 24 h, prominently in the choroid plexus. The appearance of immunoreactive T-kininogen was demonstrated in the epithelium of the choroid plexus, but not in the matrix and vessels, after i.c.v. injection of LPS. The mRNA levels of kallikreins, such as tissue kallikrein, T-kininogenase and plasma kallikrein, and B(2)-receptor did not change in any brain region following i.c.v. injection of LPS. The levels of cyclooxygenase-2 mRNA in the choroid plexus were increased within 2 h after i.c.v. injection of LPS, and pretreatment with indomethacin (3 microg/animal, i.c.v.) abolished the LPS-induced elevation of T- and H-kininogen mRNAs in the choroid plexus. The i.c.v. injection of prostaglandin E(2) (100 ng/animal) also caused increases in the mRNA levels of T- and H-kininogens in various brain regions, including the choroid plexus. These results suggest that LPS stimulates the induction of kininogens in the brain, especially the choroid plexus, by stimulating the production of arachidonic metabolites such as prostaglandin E(2).

• Down-regulation of bradykinin B2-receptor mRNA in the heart in pressure-overload cardiac hypertrophy in the rat.

  Authors: Katsutoshi Yayama, Satoshi Matsuoka, Makoto Nagaoka, Eri Shimazu, Masaoki Takano, Hiroshi Okamoto

  Biochemical pharmacology. 04/2003; 65(6):1017-25.

  To determine the potential role of the cardiac kallikrein-kinin system in the development of cardiac hypertrophy, we studied the expression patterns of kallikrein, kininogen, and bradykinin receptorTo determine the potential role of the cardiac kallikrein-kinin system in the development of cardiac hypertrophy, we studied the expression patterns of kallikrein, kininogen, and bradykinin receptor mRNA in the heart by polymerase chain reaction during the development of pressure-overload-induced left ventricular hypertrophy (LVH) in rats. The abdominal aortic constriction produced LVH after 7, 14, and 28 days. Neither mRNA levels for high-molecular-weight (H-) or low-molecular-weight (L-) kininogens and T-kininogen, nor those for tissue kallikreins, changed during LVH. B(2)-receptor mRNA levels in the left ventricles decreased 4 and 7 days after aortic constriction, subsequently returning to the levels in sham-operated animals. B(2)-receptor densities in cardiac membrane preparations obtained 4 days after aortic constriction significantly decreased compared to preparations from sham-operated rats, whereas the receptor affinity was unchanged. Down-regulation of B(2)-receptor mRNA levels was abolished by oral administration of an angiotensin II type 1 (AT1) receptor antagonist, candesartan, for 4 days after aortic constriction. Both cardiomyocytes and nonmyocytes obtained from neonatal rat hearts expressed B(2)-receptor mRNA in vitro, and the levels were not changed in either cell type by culture with 1 microM angiotensin II (Ang II). However, when a mixture of cardiomyocytes and nonmyocytes was cultured with 1 microM Ang II, B(2)-receptor mRNA levels decreased within 12 hr; this in vitro effect of Ang II was inhibited by the AT1-receptor antagonist losartan. These results indicate that the mechanical load in the myocardium caused by pressure-overload rapidly produces a down-regulation of B(2)-receptor expression during the initial stage of LVH, probably mediated by activating the AT1-receptor.

• Tissue kallikrein is synthesized and secreted by human vascular endothelial cells.

  Authors: Katsutoshi Yayama, Naomi Kunimatsu, Yumiko Teranishi, Masaoki Takano, Hiroshi Okamoto

  Biochimica et biophysica acta. 03/2003; 1593(2-3):231-8.

  The generation of kinins on the surface of vascular endothelium has been postulated in two pathways involving plasma kallikrein and tissue kallikrein; the former pathway has been well documented, butThe generation of kinins on the surface of vascular endothelium has been postulated in two pathways involving plasma kallikrein and tissue kallikrein; the former pathway has been well documented, but the latter is controversial. To clarify the presence of a kinin-generating system on endothelium, we examined whether human umbilical vein endothelial cells (HUVEC) synthesize and release tissue kallikrein in vitro. Kallikrein-like activity hydrolyzing a peptide Pro-Phe-Arg-4-methyl-coumaryl-7-amide was detected in the culture medium of HUVEC and was inhibited by aprotinin but not by soybean trypsin inhibitor. Western blotting of HUVEC medium using anti-human tissue kallikrein antibodies demonstrated the release of tissue kallikrein from HUVEC, and the reverse transcription-polymerase chain reaction (RT-PCR) followed by Southern blotting revealed the expression of tissue kallikrein mRNA in HUVEC. HUVEC metabolically labeled with [35S]methionine released radioactive proteins corresponding to tissue kallikrein. RT-PCR also showed the expression of low-molecular-weight kininogen (L-kininogen) mRNA in HUVEC. The cGMP levels in HUVEC were significantly elevated by the incubation with angiotensin converting enzyme inhibitor, lisinopril, and the elevation was completely inhibited by aprotinin or bradykinin B2-receptor antagonist, FR172357. These results suggest that the endothelial cells continuously release an active form of tissue kallikrein which enables generation of kinins on the vascular endothelium.

• Expression of kininogen mRNAs and plasma kallikrein mRNA by cultured neurons, astrocytes and meningeal cells in the rat brain

  Authors: Masaoki Takano, Masato Horie, Masanori Narahara, Masaharu Miyake, Hiroshi Okamoto

  Immunopharmacology.

  Expression of kininogen mRNAs has been studied in cultures of three different types of cells in rat brain, including neurons and astrocytes from cerebral cortex and meningeal cells from theExpression of kininogen mRNAs has been studied in cultures of three different types of cells in rat brain, including neurons and astrocytes from cerebral cortex and meningeal cells from the leptomeninges/choroid plexus. T-kininogen mRNA was expressed by meningeal cells, but not by neurons and astrocytes, and the expression in meningeal cells was enhanced by culture with prostaglandin E2 (PGE2) or dibutyryl cAMP (Bt2cAMP). Low-molecular-weight kininogen mRNA was not detected in these cultures of cells, even after treatment with PGE2. Although expression of high-molecular-weight kininogen mRNA was very low in these cultures of cells, PGE2 or Bt2cAMP markedly stimulated its expression in cultures of meningeal cells and slightly in neurons, but not in astrocytes. We also found that expression of plasma kallikrein mRNA was strong in cultures of meningeal cells and slight in astrocytes, but absent in neurons. These results suggest that cells in the leptomeninges/choroid plexus are major sources of kininogens in rat brain which may function as precursor proteins for kinins and/or potent cysteine proteinase inhibitors during cerebral inflammation.

• Expression of kininogen, kallikrein and kinin receptor genes by rat cardiomyocytes

  Authors: Katsutoshi Yayama, Makoto Nagaoka, Masaoki Takano, Hiroshi Okamoto

  Biochimica et Biophysica Acta (BBA) - Molecular Cell Research.

  To ascertain the existence of the kallikrein-kinin system in the heart, we have studied in vivo and in vitro whether rat cardiac tissue expresses kininogen, kallikrein and kinin receptor mRNAs. TheTo ascertain the existence of the kallikrein-kinin system in the heart, we have studied in vivo and in vitro whether rat cardiac tissue expresses kininogen, kallikrein and kinin receptor mRNAs. The reverse transcription-polymerase chain reaction demonstrated that the ventricular myocardium of adult male rats expressed mRNAs for T- and low-molecular-weight (L-) kininogens, tissue kallikreins such as true kallikrein and T-kininogenase, and bradykinin B2 receptor, but not those for high-molecular-weight kininogen and B1 receptor. Lipopolysaccharide (LPS; 0.5 mg/kg, i.v.) increased the levels of mRNA for T-kininogen at 12 h and the bradykinin B1 receptor at 24 h without affecting that for other components. All of these mRNAs for the kallikrein-kinin system were also detected in cultured cardiomyocytes derived from neonatal rat ventricles; dibutyryl cyclic AMP, LPS or inflammatory cytokines such as interleukin-1 and tumor necrosis factor, up-regulated mRNA expression of T-kininogen, T-kininogenase, or B1 receptor in these cells in vitro. These results suggest that there are two kinin-generating systems in rat myocardium comprising T-kininogen/T-kininogenase and L-kininogen/true kallikrein respectively, and that the former may be relatively important in inflammatory diseases or conditions in which cAMP levels increase in cardiomyocytes.