Biomedical subjects

K Furuichi

Publications and source records attributed to K Furuichi.

At least 19 recordsLinked to original sources

PubMed2006

Blockade of VEGF accelerates proteinuria, via decrease in nephrin expression in rat crescentic glomerulonephritis.

Vascular endothelial growth factor (VEGF) is a potent angiogenic factor that maintains the glomerular and peritubular capillary (PTC) network in the kidney. The soluble form of the VEGF receptor-1 (soluble fms-like tyrosine kinase 1 (sFlt-1)) is known to regulate VEGF activity by binding VEGF in the circulation. We hypothesized that VEGF may be beneficial for maintaining glomerular filtration barrier and vascular network in rats with progressive glomerulonephritis (GN). For blockade of VEGF activity in vivo, rats were transfected twice with plasmid DNA encoding the murine sFlt-1 gene into femoral muscle 3 days before and 2 weeks after the induction of antiglomerular basement membrane antibody-induced GN. Inhibition of VEGF with sFlt-1 resulted in massive urinary protein excretion, concomitantly with downregulated expression of nephrin in nephritic rats. Further, blockade of VEGF induced mild proteinuria in normal rats. Administration of sFlt-1 affected neither the infiltration of macrophages nor crescentic formation. In contrast, treatment of sFlt-1 accelerated the progression of glomerulosclerosis and interstitial fibrosis accompanied with renal dysfunction and PTC loss at day 56. VEGF may play a role in maintaining the podocyte function as well as renal vasculature, thereby protecting glomeruli and interstitium from progressive renal insults.

Animals↗

PubMed2005

The light-addressable potentiometric sensor for multi-ion sensing and imaging.

The light-addressable potentiometric sensor (LAPS) is a semiconductor-based chemical sensor with an electrolyte-insulator-semiconductor structure. The LAPS can have many measuring points integrated on the sensing surface, which are individually accessed by a light beam. By modifying the measuring points with different materials, a single sensor plate can be used as a multi-analyte sensor. In this paper, instrumentation and application of LAPS to multi-ion sensing and imaging are described. As a new application of LAPS, potentiometric imaging of a microfluidic channel is proposed.

Biosensing Techniques↗

PubMed2003

Response to ADP-ribose by activation of TRPM2 in the CRI-G1 insulinoma cell line.

The response to intracellular ADP-ribose in the rat CRI-G1 insulinoma cell line was studied using a patch-clamp method. Dialysis of ADP-ribose into cells induced a response in a dose-dependent manner. The reversal potentials in various solutions showed that the ADP-ribose-gated channel was a Ca2+-permeable nonselective cation channel. In inside-out recordings, ADP-ribose and b-NAD induced responses in the same patch. The single-channel current-voltage relationships for ADP-ribose- and b-NAD-induced responses were almost identical, indicating that ADP-ribose and b-NAD activated the same channel. The physiological properties of the ADP-ribose-gated channel are similar to those we reported previously for the cloned transient receptor potential channel TRPM2. Moreover, RT-PCR analysis showed that TRPM2 was abundantly expressed in CRI-G1 cells, suggesting that the ADP-ribose-gated channel represents the native TRPM2 channel in CRI-G1 cells. These results suggest that ADP-ribose can be an endogenous modulator of Ca2+ influx through the TRPM2 channel into CRI-G1 cells.

Adenosine Diphosphate Ribose↗

PubMed2001

Functional characterization of cysteinyl leukotriene CysLT(2) receptor on human coronary artery smooth muscle cells.

Cysteinyl leukotrienes (LTC(4), LTD(4), and LTE(4)) are a class of biologically active lipids that exert potent effects on the heart. To assess their roles, we investigated the distribution of their receptors, CysLT(1) and CysLT(2), in the cardiovascular system. CysLT(2) mRNA was detected at high levels in the human atrium and ventricle and at intermediate levels in the coronary artery, whereas CysLT(1) mRNA was barely detected. Further analysis by in situ hybridization revealed that CysLT(2) mRNA was expressed in myocytes, fibroblasts, and vascular smooth muscle cells, but not in endothelial cells. When human coronary smooth muscle cells were stimulated with LTC(4), the intracellular calcium concentration increased in a dose-dependent manner, and this action was partially inhibited by nicardipine. Additionally, these cells showed chemotactic responses to LTC(4). This is the first report on the physiological role of CysLT(2), and the findings suggest that CysLT(2) has biological significance in the cardiovascular system.

Arteries↗

PubMed2001

Immunocyte Ca2+ influx system mediated by LTRPC2.

We characterized an activation mechanism of the human LTRPC2 protein, a member of the transient receptor potential family of ion channels, and demonstrated that LTRPC2 mediates Ca2+ influx into immunocytes. Intracellular pyrimidine nucleotides, adenosine 5'-diphosphoribose (ADPR), and nicotinamide adenine dinucleotide (NAD), directly activated LTRPC2, which functioned as a Ca2+-permeable nonselective cation channel and enabled Ca2+ influx into cells. This activation was suppressed by intracellular adenosine triphosphate. These results reveal that ADPR and NAD act as intracellular messengers and may have an important role in Ca2+ influx by activating LTRPC2 in immunocytes.

ADP-ribosyl Cyclase↗

PubMed2001

Involvement of p38 mitogen-activated protein kinase followed by chemokine expression in crescentic glomerulonephritis.

p38 Mitogen-activated protein kinase (MAPK) is involved in the production and signal transduction of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha, and chemokines in vitro. However, the crucial role of p38 MAPK in the inflammatory processes of crescentic glomerulonephritis in vivo remains to be investigated. We showed a dramatic decrease in IL-1beta-induced phosphorylation of p38 MAPK, not extracellular signal-regulated kinases 1/2 or jun NH2-terminal kinase, in rat cultured mesangial cells by FR167653. We explored the effects of FR167653 as a specific inhibitor of p38 MAPK on renal injury and subsequent renal expression of chemokines in a progressive experimental crescentic glomerulonephritis model in Wistar-Kyoto rats. Rats developed crescentic glomerulonephritis leading to glomerulosclerosis and interstitial fibrosis by 56 days after the administration of nephrotoxic sera. The number of phosphorylated p38 MAPK-positive cells, detected mainly in crescents, correlated well with the percentage of crescents and number of ED-1-positive cells. Phosphorylated p38 MAPK-positive cells were downregulated in glomeruli in rats with the daily subcutaneous administration of FR167653 for 6 days. Concomitantly, renal expression of macrophage inflammatory protein-1alpha and monocyte chemoattractant protein-1/monocyte chemotactic and activating factor was markedly reduced by day 6. The severity of glomerulosclerosis and interstitial fibrosis significantly decreased by day 56, and renal function was preserved. These results suggest that p38 MAPK phosphorylation is pivotal for crescentic glomerulonephritis, followed by the subsequent expression of renal chemokines. This study provides evidence that regulation of p38 MAPK is a novel appealing therapeutic target for crescentic glomerulonephritis.

Animals↗

PubMed2001

Upregulation of fractalkine in human crescentic glomerulonephritis.

BACKGROUND/AIM: To evaluate the importance of fractalkine, a novel member of the CX3C chemokine, and natural killer (NK) cells in human crescentic glomerulonephritis, we determined the presence of fractalkine in the diseased kidneys immunohistochemically, and the correlation among fractalkine, NK cells and the degree of renal damage. METHODS: Twenty-three patients (13 males and 10 females) with primary or secondary crescentic glomerular disease were evaluated in this study. Fractalkine and CD16-positive cells including NK cells were detected immunohistochemically. RESULTS: Fractalkine-positive cells were detected in the interstitium of 23 patients with crescentic glomerulonephritis, while they were not detected in the glomeruli. In addition, CD16-positive cells were detected in both the glomeruli (1.3 +/- 0.2/glomerulus) and interstitium (1.3 +/- 0.2/visual field). The number of fractalkine-positive cells in the interstitium correlated with the number of CD16-positive cells before glucocorticoid therapy (r = 0.43, p = 0.047, n = 23). The number of fractalkine-positive cells in the interstitium before glucocorticoid therapy (0.2 +/- 0.1/visual field) decreased after therapy (0.1 +/- 0.1/visual field, p = 0.050) in 11 cases tested. The number of CD16-positive cells in the diseased kidneys did not change after glucocorticoid therapy. CONCLUSION: These results suggest that the local production of fractalkine may explain the presence of CD16-positive cells including NK cells, which may participate in the interstitial lesions of human crescentic glomerulonephritis before corticoid therapy.

Adult↗

PubMed2001

Serum levels of KL-6 reflect disease activity of interstitial pneumonia associated with ANCA-related vasculitis.

OBJECTIVE: KL-6 is reported to be excreted from the lung alveolar and bronchial epithelial cells and may be a good marker for monitoring disease activity of interstitial pneumonia. This study was designed to ascertain the clinical significance of serum KL-6 levels in interstitial pneumonia associated with anti-neutrophil cytoplasmic antibody (ANCA)-related vasculitis. METHODS: Serum KL-6 levels were determined by an enzyme-linked immunosorbent assay. PATIENTS: We examined 20 healthy subjects, 13 patients with perinuclear (myeloperoxidase, MPO) ANCA-related vasculitis and 12 dermatomyositis (DM)/polymyositis (PM) patients as disease controls in this study. Six out of 13 patients with ANCA-related vasculitis had interstitial pneumonia. RESULTS: Serum levels of KL-6 in ANCA-positive patients with interstitial pneumonia were significantly elevated, while they remained as low as those of healthy subjects in ANCA-positive patients without interstitial pneumonia. Similarly, KL-6 levels in sera were higher in 12 dermatomyositis/polymyositis patients with interstitial pneumonia, while they remained low in DM/PM patients without interstitial pneumonia. Moreover, the elevated serum KL-6 level was reduced during the convalescence induced by glucocorticoid therapy and reflected the disease activity of interstitial pneumonia associated with ANCA-related vasculitis. CONCLUSION: These data suggest that the measurement of serum KL-6 levels may be a good monitoring system for the diagnosis and follow-up of interstitial pneumonia of patients with ANCA-related vasculitis.

Aged↗

PubMed2001

[Effect of interferon therapy on hepatitis B virus related membranous nephropathy associated with focal segmental glomerulosclerosis].

We experienced a 24-year-old Japanese man, who was a hepatitis B virus carrier with nephrotic syndrome. Liver biopsy showed that he was suffering from chronic hepatitis (activity 2, fibrosis 2). Renal biopsy revealed membranous nephropathy(MN) with focal segmental glomerulosclerosis(FGS). Immunofluorescentic findings revealed the presence of HBe antigen along the glomerular capillaries as well as HBe antigenemia in circulation. Therefore, we diagnosed this case as HB virus-related membranous nephropathy associated with FGS lesions. He was treated with interferon(IFN) alpha-2b for over a month and angiotensin converting enzyme inhibitor. These therapies reduced urinary protein excretion from 4-6 g/day to 1-2 g/day, in accordance with a decrease in the titer of HBV DNA polymerase. The second renal biopsy revealed that the histological change from MN to membranoproliferative glomerulonephritis Type III after IFN therapy. These results suggest that IFN therapy might be effective for HB virus-related MN associated with FGS.

Adult↗

PubMed2001

Molecular cloning of the platelet P2T(AC) ADP receptor: pharmacological comparison with another ADP receptor, the P2Y(1) receptor.

Platelet activation plays an essential role in thrombosis. ADP-induced platelet aggregation is mediated by two distinct G protein-coupled ADP receptors, Gq-linked P2Y(1), and Gi-linked P2T(AC), which has not been cloned. The cDNA encoding a novel G protein-coupled receptor, termed HORK3, was isolated. The HORK3 gene and P2Y(1) gene were mapped to chromosome 3q21-q25. HORK3, when transfected in the rat glioma cell subline (C6-15), responded to 2-methylthio-ADP (2MeSADP) (EC(50) = 0.08 nM) and ADP (EC(50) = 42 nM) with inhibition of forskolin-stimulated cAMP accumulation. 2MeSADP (EC(50) = 1.3 nM) and ADP (EC(50) = 18 nM) also induced intracellular calcium mobilization in P2Y(1)-expressing cells. These results show that HORK3 is a Gi/o-coupled receptor and that its natural ligand is ADP. AR-C69931 MX and 2MeSAMP, P2T(AC) antagonists, selectively inhibited 2MeSADP-induced adenylyl cyclase inhibition in HORK3-expressing cells. On the other hand, A3P5PS, a P2Y(1) antagonist, blocked only 2MeSADP-induced calcium mobilization in P2Y(1)-expressing cells. HORK3 mRNA was detected in human platelets and the expression level of HORK3 was equivalent to that of P2Y(1). These observations indicate that HORK3 has the characteristics of the proposed P2T(AC) receptor. We have also determined that [(3)H]2MeSADP binds to cloned HORK3 and P2Y(1). Competition binding experiments revealed a similarity in the rank orders of potency of agonists and the selectivity of antagonists as obtained in the functional assay. These results support the view that P2Y(1) functions as a high-affinity ADP receptor and P2T(AC) as a low-affinity ADP receptor in platelets.

Adenosine Triphosphate↗

PubMed2000

Molecular cloning and characterization of another leukotriene B4 receptor.

Leukotriene B(4) is a potent lipid mediator known to be implicated mainly in inflammatory actions. Previous pharmacological studies indicated the existence of only one class of G protein-coupled receptor for leukotriene B(4), for which a candidate gene, namely BLT, had been identified. Here we report the isolation of another gene encoding a functional G protein-coupled receptor for leukotriene B(4), named JULF2. JULF2 is a novel G protein-coupled receptor of 358 amino acids that shares 36.6% amino acid identity with human BLT. According to genomic information, the JULF2 gene is located on the chromosome 14, about 4 kilobases upstream of the BLT gene. During screening of endogenous ligands for JULF2, we found that leukotriene B(4) induced inhibition of forskolin-stimulated cAMP accumulation in Chinese hamster ovary cells, stably expressing JULF2. Additionally, Chinese hamster ovary cells expressing exogenous JULF2 showed chemotactic responses with leukotriene B(4) in a pertussis toxin-sensitive manner. A large amount of JULF2 mRNA was detected in the human spleen and the peripheral blood leukocytes. Furthermore, JULF2 mRNA was expressed in mononuclear lymphocytes, in which BLT mRNA was barely detected. The discovery of this second leukotriene B(4) receptor will eventually lead to a better understanding of the classification of leukotriene B(4) receptors and reconsideration of the pathophysiological role of leukotriene B(4).

Amino Acid Sequence↗

PubMed2000

Evidence that atypical vasopressin V(2) receptor in inner medulla of kidney is V(1B) receptor.

Vasopressin V(2) receptors at high-density and V(1B) receptors are candidates for the V(2)-like receptor, which evokes an increase in [Ca(2+)](i) when stimulated by the vasopressin V(2) receptor agonist 1-desamino-8-D-arginine vasopressin (dDAVP) in kidney inner medullary collecting duct. We compared the pharmacological characteristics of vasopressin V(2) and V(1B) receptors in Chinese hamster ovary (CHO) cells to those of vasopressin V(2)-like receptors in rat inner medullary collecting duct cells. The vasopressin V(1B) receptor-selective agonist [deamino-Cys(1), D-3-(Pyridyl)-Ala(2), Arg(8)]vasopressin (D3PVP) did not stimulate the [Ca(2+)](i) increase in high-density vasopressin V(2) receptor-expressing CHO cells, but did in inner medullary collecting duct cells. Moreover, the vasopressin V(1A)/V(2) receptor dual antagonist 4'-[(2-methyl-1,4,5,6-tetrahydroimidazo[4,5-d][1] benzazepin-6-yl)carbonyl] 2-phenylbenzanilide (YM087), which has no effect on vasopressin V(1B) receptors, did not block the [Ca(2+)](i) increase in inner medullary collecting duct cells when stimulated by dDAVP and D3PVP. On reverse transcription-polymerase chain reaction (RT-PCR) analysis of kidney, vasopressin V(1B) receptor mRNA was detected only in the medulla. We propose that the true nature of the vasopressin V(2)-like receptor in the inner medullary collecting duct is the vasopressin V(1B) receptor, rather than the vasopressin V(2) receptor expressed at high-density.

Animals↗

PubMed2000

The molecular characterization and tissue distribution of the human cysteinyl leukotriene CysLT(2) receptor.

Cysteinyl leukotrienes (CysLTs), slow-reacting substances of anaphylaxis, are lipid mediators known to possess potent proinflammatory action. Pharmacological studies using CysLTs indicate that at least two classes of G protein-coupled receptors (GPCRs), named CysLT(1) and CysLT(2), exist; the former is sensitive and the latter is resistant to the CysLT(1) antagonists currently used to treat asthma. Although the CysLT(1) receptor gene has been recently cloned, the molecular identity of the CysLT(2) receptor has remained elusive. Here we show that the pharmacological profile of an orphan GPCR (PSEC0146) is consistent with that of the CysLT(2) receptor. In human embryonic kidney 293 cells that express the PSEC0146 cDNA, leukotriene C(4) (LTC(4)) and leukotriene D(4) (LTD(4)) induce equal increases in intracellular calcium mobilization; these increases are not affected by CysLT(1) antagonists. Additionally, [(3)H]LTC(4) specifically binds to membranes from COS-1 cells transiently transfected with PSEC0146. Large amounts of the PSEC0146 mRNA are found in human heart, placenta, spleen, and peripheral blood leukocytes but not in the lung and the trachea. Pharmacological feature and expression studies will eventually lead to a better understanding of the classification of CysLT receptors, possibly leading to a reconsideration of the pathological and physiological role of CysLTs.

Animals↗

PubMed2000

Cloning, expression, and characterization of ferret 5-HT(3) receptor subunit.

Ferrets (Mustela putorius furo) are useful animals for determining anti-emetic activity via 5-HT(3) receptors in vivo. We isolated a cDNA encoding the 5-hydroxytryptamine (5-HT) type 3A receptor subunit (5-HT(3A)) from ferret colon, expressed it in a human embryonic kidney cell line and determined its pharmacological properties. The open reading frame of the isolated cDNA encoded a 483-amino acid protein, corresponding to the shorter splice variant of 5-HT(3A) receptors. Splice variants were no longer detected by reverse transcriptase-polymerase chain reaction. The ferret 5-HT(3A) receptor exhibits a high degree of amino acid sequence identity (>/=80%) to that of other species. Binding studies demonstrated the following rank order of potency for agonists: meta-chlorophenylbiguanide (mCPBG)>2-methyl-5-hydroxytryptamine (2-Me-5-HT)=5-HT, and for antagonists: ondansetron=tropisetron>(+)-tubocurarine>metoclopramide. Electrophysiological studies revealed that mCPBG was a partial agonist and 2-Me-5-HT was an almost fully effective agonist compared to 5-HT.

Alternative Splicing↗

PubMed2000

An evolutionarily conserved G-protein coupled receptor family, SREB, expressed in the central nervous system.

We report here a novel family of G-protein coupled receptor (GPCR) which is extraordinarily conserved among vertebrate species. This family, designated SREB (Super Conserved Receptor Expressed in Brain), consists of at least three members, termed SREB1, SREB2, and SREB3. SREB members share 52-63% amino acid identity with each other and show relatively high similarity to previously known amine amine GPCRs (approximately 25% identity). Amino acid sequence identity between human and rat orthologues is 97% for SREB1 and 99% for SREB3, while the SREB2 sequence is surprisingly completely identical between the species. Furthermore, amino acid sequence of zebrafish SREB2 and SREB3 are 94 and 78% identical to mammal orthologues. Northern blot analysis revealed that SREB members are predominantly expressed in the brain regions and genital organs. Radiation hybrid analysis localized SREB1, SREB2, and SREB3 genes to different human chromosomes, namely 3p21-p14, 7q31 and Xp11, respectively. The high sequence conservation and abundant expression in the central nervous system suggest the existence of undiscovered fundamental neuronal systems consisting of SREB family members and their endogenous ligand(s).

Amino Acid Sequence↗

PubMed2000

The novel G-protein coupled receptor SALPR shares sequence similarity with somatostatin and angiotensin receptors.

A cDNA encoding a novel G-protein coupled receptor (GPCR) was isolated from a human cerebral cortex cDNA library by low stringency hybridization screening. This putative seven-transmembrane domain receptor of 469 amino acids was designated SALPR (Somatostatin- and Angiotensin- Like Peptide Receptor). SALPR shares the highest amount of amino acid similarity with the somatostatin (35% with SSTR5) and angiotensin receptors (31% with AT1). Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that the SALPR mRNA is predominantly expressed in human brain regions, particularly the substantia nigra and pituitary, although the mRNA can also be detected in the peripheral tissues, albeit at low levels. Chromosomal mapping by radiation hybrid analysis localized the human SALPR gene to the chromosome 5p15.1-5p14. Transient expression of SALPR in COS-1 cells did not produce any binding sites for somatostatin or angiotensin II, indicating the necessity for further study to discover its ligand and physiological significance.

Amino Acid Sequence↗

PubMed2000

Homodimer of two F-box proteins betaTrCP1 or betaTrCP2 binds to IkappaBalpha for signal-dependent ubiquitination.

We found previously that overexpression of an F-box protein betaTrCP1 and the structurally related betaTrCP2 augments ubiquitination of phosphorylated IkappaBalpha (pIkappaBalpha) induced by tumor necrosis factor-alpha (TNF-alpha), but the relationship of the two homologous betaTrCP proteins remains unknown. Herein we reveal that deletion mutants of betaTrCP1 and betaTrCP2 lacking the F-box domain suppressed ubiquitination and destruction of pIkappaBalpha as well as transcriptional activation of NF-kappaB. The ectopically expressed betaTrCP1 and betaTrCP2 formed both homodimer and heterodimer complexes without displaying the trimer complex. Dimerization of betaTrCP1 and/or betaTrCP2 takes place at their conserved NH(2)-terminal regions, termed a "D-domain" (for dimerization domain), located upstream of the F-box domain. The D-domain was necessary and sufficient for the dimer formation. Intriguingly, the betaTrCP homodimer, but not the heterodimer, was selectively recruited to pIkappaBalpha induced by TNF-alpha. These results indicate that not only betaTrCP1 but also betaTrCP2 participates in the ubiquitination-dependent destruction of IkappaBalpha by forming SCF(betaTrCP1-betaTrCP1) and SCF(betaTrCP2-betaTrCP2) ubiquitin-ligase complexes.

Base Sequence↗

PubMed2000

Up-regulation of monocyte chemoattractant protein-1 in tubulointerstitial lesions of human diabetic nephropathy.

BACKGROUND: We previously described that monocyte chemoattractant protein-1 (MCP-1) plays an important role in progressive glomerular and interstitial damage in inflammatory renal diseases. However, the expression of MCP-1 in diabetic nephropathy remains to be investigated. METHODS: We examined whether locally expressed MCP-1 participates in human diabetic nephropathy via recruiting and activating monocytes/macrophages (Mphi). Urinary and serum MCP-1 levels were measured by enzyme-linked immunosorbent assay in 45 patients with diabetic nephropathy. The presence of MCP-1 in diseased kidneys was determined by immunohistochemical and in situ hybridization analyses. RESULTS: Urinary MCP-1 levels were significantly elevated in patients with diabetic nephrotic syndrome and advanced tubulointerstitial lesions. Moreover, urinary levels of MCP-1 were well correlated with the number of CD68-positive infiltrating cells in the interstitium. In contrast, serum MCP-1 levels remained similar to those of healthy volunteers. Furthermore, we detected the MCP-1-positive cells in the interstitium of diabetic nephropathy via both immunohistochemical and in situ hybridization analyses. CONCLUSION: These observations suggest that locally produced MCP-1 may be involved in the development of advanced diabetic nephropathy, especially in the formation of tubulointerstitial lesions possibly through Mphi recruitment and activation. Moreover, up-regulation of MCP-1 may be a common pathway involved in the progressive tubulointerstitial damage in diabetic nephropathy as well as inflammatory renal diseases.

Adolescent↗