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The occurrence of more than two rearranged bands of immunoglobulin heavy chain (IgH) genes in B precursor acute lymphoblastic leukemia (ALL) has recently been documented. To elucidate the nature of such leukemias, we studied 30 patients with common ALL, including 6 patients with Philadelphia chromosome (Ph1)-positive ALL, by immunophenotyping and genotyping. In 10 of the 30, Southern blotting showed oligoclonal patterns of IgH gene arrangements, which were frequently detected in Ph1-positive ALL. In one patient of the 10, three rearranged bands of Ig kappa chain genes were detected. Ph1 abnormality and co-expression of myeloid associated antigens were found in 5 and 5 of the 10, respectively. Detection of multiple fragments of IgH genes would be suggestive of multipotent progenitor origin of these ALL.

Recent advances in technologies such as DNA microarrays have provided an abundance of gene expression data on the genomic scale. One of the most important projects in the post-genome-era is the systemic identification of gene expression networks. However, inferring internal gene expression structure from experimentally observed time-series data are an inverse problem. We have therefore developed a system for inferring network candidates based on experimental observations. Moreover, we have proposed an analytical method for extracting common core binomial genetic interactions from various network candidates. Common core binomial genetic interactions are reliable interactions with a higher possibility of existence, and are important for understanding the dynamic behavior of gene expression networks. Here, we discuss an efficient method for inferring genetic interactions that combines a Step-by-step strategy (Y. Maki, Y. Takahashi, Y. Arikawa, S. Watanabe, K. Aoshima, Y. Eguchi, T. Ueda, S. Aburatani, S. Kuhara, M. Okamoto, An integrated comprehensive workbench for inferring genetic networks: Voyagene, Journal of Bioinformatics and Computational Biology 2(3) (2004) 533.) with an analysis method for extracting common core binomial genetic interactions.

Pneumonia is a critical issue during the agonal phase, and often becomes lethal in the absence of pathogen detection. Autopsy is a powerful tool for analyzing the cause of a patient's death, progression of the disease, and the therapeutic response. However, it is frequently limited to the identification of bacterial strains. To elucidate the pathogenesis during the agonal phase of pneumonia, intrapulmonary sputum was harvested by directly inserting a swab into a resected lung, and the bacterial composition was analyzed using both pathological and microbiological techniques from 15 patients with hematological malignancies, and the results were compared with those from 25 patients with other medical and surgical diseases. Among the 54 bacteria strains isolated from the 40 patients, multidrug-resistant strains were significantly more prevalent in hematological group than in other diseases (16/21 versus 11/33, P = .002). Enterococcus faecium was preferentially isolated from the hematological patients, whereas the methicillin-resistant Staphylococcus aureus was predominantly found in the nonhematological group. Two coagulase-negative Staphylococcus epidermidis strains in hematological diseases may be diagnosed as causative bacteria of pneumonia by both bacterial and pathological techniques. Although the results of this study may not be directly applicable for clinical diagnosis, this approach has a potential to become not only a diagnostic method for bacterial pneumonia, but may be also useful for the analysis of multidrug-resistant pathogens.

Low-dose cytarabine (ara-C) has been used to treat older patients with acute myeloid leukemia (AML) or high-risk myelodysplastic syndrome (MDS), but has resulted in complete remission for20% of cases. A pilot study of the efficacy of a combination chemotherapy using low-dose ara-C, melphalan (Mel), and mitoxantrone (Mit) was conducted.The treatment comprised ara-C (10 mg/m2) twice daily, melphalan (2 mg/body) every other day, and mitoxantrone (3 mg/m2) every 3 days. The treatment was discontinued if the nuclear cell count was15,000/microl with20% blast count in the bone marrow. The primary end-points were initial response and tolerability.The study comprised 9 patients with AML or high-risk MDS (median age, 75 years). Complete remission was achieved in 3 patients. All the patients displayed grade 4 neutropenia and thrombocytopenia. One patient died from sepsis.The present regimen was more effective and displayed similar safety, compared with low-dose ara-C alone.

DNA fragmentation into internucleosomal fragments is the best recognized biochemical event of apoptosis. Two major caspase pathways have been identified in the signal transduction leading to DNA fragmentation: the receptor pathway and the mitochondrial pathway. DNA fragmentation factor (DFF) has been identified as a major apoptotic endonuclease in the internucleosomal DNA fragmentation process. However, the potential roles of caspases and DFF in internucleosomal DNA fragmentation induced by specific stimuli still need to be investigated since caspase-independent pathways and nuclease(s) other than DFF also play important roles during this process. In the present study, we investigated the activity of GP7 (4-[4 ''-(2 '',2 '',6 '',6 ''-tetramethyl-1 ''-piperidinyloxy) amino]-4'-demethyl epipodophyllotoxin), a new spin-labeled derivative of podophyllotoxin semi-synthesized by our university, to induce apoptosis of the human leukemia cell line NB4. GP7 induced the release of cytochrome-c from mitochondria, activations of caspase-3, -8, and -9, cleavage of DFF45/inhibitor of caspase-activated DNase, activation of DFF40/caspase-activated DNase, and apoptotic DNA fragmentation in NB4 cells. The broad-spectrum caspase inhibitor zVAD-fmk abrogated GP7-induced caspase-3, -8, and -9 activations but could not inhibit GP7-induced apoptotic DNA fragmentation in NB4 cells. Our findings suggest that GP7-induced apoptotic DNA fragmentation in NB4 cells is independent of caspase activation and DFF, although they are closely involved in this process.

Background Angiotensin-converting enzyme (ACE) inhibitors are widely used in treatment of heart failure, but little is known regarding whether ACE inhibitors regulate the activity of matrix metalloproteinases (MMPs) and the tissue inhibitor of MMPs (TIMPs) in cardiac cells. The purpose of this study was to determine the ability and possible signal pathway involved of imidaprilat, an ACE inhibitor, to modulate MMP-2 and TIMP-2 in human cardiac fibroblasts in the presence of interleukin (IL)-1β. Methods and results Using gelatin zymography and RT-PCR and Griess analysis,we found that IL-1β increased the MMP-2 activity and transcription and nitric oxide(NO) production from supernatant of culture medium.These effects of IL-1β were inhibited by imidaprilat or the NO synthase inhibitor, L-NMMA. Sodium nitroprusside (SNP), an exogenous NO donor, prevented significantly the effects of imidaprilat on MMP-2 inhibition. Imidaprilat alone didn't affect MMP-2 activity and expression. Neither IL-1β nor imidaprilat has no effect on TIMP-2 transcription in cardiac fibroblasts. Conclusions The current study demonstrates imidaprilat inhibits MMP-2 activity and expression in human cardiac fibroblasts induced by IL-1β via NO-dependent pathway. These data suggest that the beneficial effect of ACE inhibitors against left cardiac remodeling and heart failure may be due at least in part to regulating MMPs activity and expression by modulation of NO pathway.

In this paper, we propose a new system extracting potentially copyright infringement texts from the Web, called EPCI. EPCI extracts them in the following way: (1) generating a set of queries based on a given copyright reserved seed-text, (2) putting every query to search engine API, (3) gathering the search result Web pages from high ranking until the similarity between the given seed-text and the search result pages becomes less than a given threshold value, and (4) merging all the gathered pages, then re-ranking them in the order of their similarity. Our experimental result using 40 seed-texts shows that EPCI is able to extract 132 potentially copyright infringement Web pages per a given copyright reserved seed-text with 94% precision in average.

Objective To study the effects of heparin on the production of homocysteine-induced extracellular matrix metalloproteinase-2 (MMP-2) in cultured rat vascular smooth muscle cells. Methods The effects of different homocysteine levels (0 μmol/L to 1000 μmol/L) on MMP-2 production and the effects of different heparin concentrations (0 μg/mL to 100 μg/mL) on homocysteineinduced MMP-2 in cultured rat vascular smooth muscle cells were examined using gelatin zymography and Western blotting. The changes in MMP-2 were further compared with various treatments for 24 h, 48 h and 72 h. Results Homocysteine (50 μmol/L to 1000 μmol/L) increased the production of MMP-2 significantly in a dose-dependent manner. Increased production of MMP-2 induced by homocysteine was reduced by the extracellular addition of heparin in a dose-dependent manner. Production of MMP-2 with various treatment regimens for 72 h was greater than for 24 h and 48 h. Conclusions Extracellular addition of heparin decreased homocysteine-induced MMP-2 secretion. Data suggest a mechanism by which hyperhomocysteinemia is involved in the pathogenesis of coronary artery disease and demonstrate a beneficial effect of heparin on these conditions.

We quantitatively assessed the expression of cytokine receptors (interleukin-2 receptor (IL-2R), IL-3R, IL-4R, IL-5R, IL-6R, IL-7R, granulocyte-macrophage colony-stimulating factor R (GM-CSFR), G-CSFR, c-fms, c-mpl, c-kit and FLT3) in cells from 211 adults with acute lymphoblastic leukemia (ALL) by flow cytometry and determined their prevalence and clinical significance. Although all cytokine receptors were expressed to various degrees, the levels of IL-3R alpha-chain (IL-3Ralpha), IL-2Ralpha, IL-2Rbeta, IL-7Ralpha, common-Rgamma(gammac), c-mpl, c-kit and FLT3 exhibited a wide spectrum > or =2000 sites/cell. Among them, IL-3Ralpha, IL-2Ralpha and FLT3 were highly expressed in B-lineage ALL, whereas IL-7Ralpha, gammac and c-kit predominated in T-lineage ALL. Higher levels of IL-3Ralpha, IL-2Ralpha, c-kit and FLT3 correlated with the expression of CD13/33. Increased IL-2Ralpha levels related to the presence of Philadelphia chromosome (Ph), leukocytosis and shorter event-free survival (EFS). C-kit preferred in male. Elevated FLT3 levels correlated with age > or =60 years. Multivariate analysis in B-lineage ALL revealed only IL-2Ralpha (P=0.028) and Ph (P=0.020) as independent factors for EFS. These findings suggest that several cytokine receptors associated with certain cellular and clinical features, but IL-2Ralpha solely had a prognostic value and should be considered as a major prognostic factor for adult ALL that is comparable with Ph.

Multiple myeloma (MM) is a molecularly heterogeneous hematologic malignancy and remains mostly incurable despite the recent improvement of treatment strategies by several novel agents. Therefore, it is important to develop more efficacious drug against MM. YM155, a novel small molecule suppressant of survivin, shows anti-proliferative activities against various human cancer cells. YM155 was identified in a survivin gene promoter assay by high throughput screening of chemical libraries. In the present study, we investigated the cytotoxic mechanism of YM155 against human myeloma cells including bortezomib (BTZ) resistant cells (U266/BTZ). Three myeloma cell lines, U266, KMS-11 and KMS-12, were employed. YM155 inhibited the cell growth of these cell lines with the IC50 value of below 5 nM. YM155 suppressed the expression of mRNA and protein of survivin. We also found that YM155 inhibited the protein expression of Mcl-1, as an essential anti-apoptotic protein for survival of myeloma cells. In addition, we observed that YM155 markedly suppressed the phosphorylation of STAT3, which is known as transcription factor of Mcl-1. When KMS-12 cells were incubated with IL-6, phosphorylation of STAT3 and upregulation of Mcl-1 protein were observed. Treatment of KMS-12 with YM155 inhibited these events and eventually induced apoptosis in KMS-12 cells. Interestingly, inhibitory effect of YM155 on Mcl-1 protein expression was much stronger than that on survivin. RQ-PCR analysis indicated that the level of Mcl-1 mRNA was not affected after YM155 treatment. Immunoblot analysis showed that proteasome inhibitor MG-132 blocked the inhibition of Mcl-1 expression by YM155, suggesting that proteasome-mediated degradation is involved in YM155-induced Mcl-1 downregulation. MM is a low-growth-fraction disease and low proliferation of MM seems to contribute to resistance to various anticancer drugs. To determine whether YM155 shows cytotoxic effect against quiescent (G0/G1) MM cells, U266 were cultured in low-serum medium to enrich the G0/G1 population. Dual-parametric flow cytometric analysis using Hoechest33342 and the RNA specific dye pyronin Y revealed that YM155 potently induced cell death of quiescent (G0/G1) MM cells. In quiescent MM cells, inhibitory effect of YM155 on Mcl-1 protein expression was much stronger than that on survivin. We also examined whether similar effect of YM155 could be observed in primary MM cells. The majority of primary MM cells from patients was found to be in quiescent phase by cell-cycle analysis. YM155 showed similar cytotoxic activity against primary MM cells. In contrast, Ara-C, the S-phase specific anticancer drug, never killed quiescent primary MM cells. We established BTZ-resistant MM cell line (U266/BTZ). The IC50 value was 45-fold higher than its parental cell line. DNA sequencing data indicated that U266/BTZ cells possess a point mutation, G322A, in the gene encoding the proteasome beta-5 subunit. YM155 almost equally exhibited cytotoxic activity against U266/BTZ compared with parental cells. U266/BTZ displayed significantly lowered amounts of bcl-2, survivin and aurora-B kinase proteins. Interestingly, U266/BTZ overexpressed the Mcl-1 protein. Treatment with YM155 rapidly suppressed Mcl-1 protein expression and induced apoptosis. These data suggest that overexpression of Mcl-1 may contribute to bortezomib resistance and downregulation of Mcl-1 by YM155 could overcome it. In conclusion, our data indicate that YM155 may exert robust cytotoxic activity against quiescent (G0/G1) MM and bortezomib resistant cells via inhibition of Mcl-1 and survivin. Disclosures No relevant conflicts of interest to declare.

A case of a lost guide wire extending from the vena cava to the back of the neck after central venous catheterization is presented. A trainee inserted a central venous catheter via the left subclavian vein in a 40-year-old male patient after surgery, but did not notice that a guide wire was completely inserted in the vein. After 6 months, the lost guide wire was seen extending from the saphenous vein through the vena cava, right atrium, right ventricle, pulmonary artery and lung tissue to the back of neck. Although percutaneous catheterization of central veins is a routine technique, it is a procedure requiring advanced surgical skills, expert supervision, and attention to detail in order to prevent adverse effects. The present case is not only a technological problem, but also one of responsibility. The operator must hold onto the guide wire at all times until removal from the vessel, and a supervisor must make sure that trainees are aware of all possible complications.

Central nervous system (CNS) relapse accompanying the prolonged administration of imatinib mesylate has recently become apparent as an impediment to the therapy of Philadelphia chromosome–positive (Ph+) leukemia. CNS relapse may be explained by limited penetration of imatinib mesylate into the cerebrospinal fluid because of the presence of P-glycoprotein at the blood-brain barrier. To overcome imatinib mesylate–resistance mechanisms such as bcr-abl amplification, mutations within the ABL kinase domain, and activation of Lyn, we developed a dual BCR-ABL/Lyn inhibitor, INNO-406 (formerly NS-187), which is 25 to 55 times more potent than imatinib mesylate in vitro and at least 10 times more potent in vivo. The aim of this study was to investigate the efficacy of INNO-406 in treating CNS Ph+ leukemia. We found that INNO-406, like imatinib mesylate, is a substrate for P-glycoprotein. The concentrations of INNO-406 in the CNS were about 10% of those in the plasma. However, this residual concentration was enough to inhibit the growth of Ph+ leukemic cells which expressed not only wild-type but also mutated BCR-ABL in the murine CNS. Furthermore, cyclosporine A, a P-glycoprotein inhibitor, augmented the in vivo activity of INNO-406 against CNS Ph+ leukemia. These findings indicate that INNO-406 is a promising agent for the treatment of CNS Ph+ leukemia.

Bcl-2 protein plays a critical role in inhibiting anticancer drug–induced apoptosis. We found that Bcl-2 overexpression is associated with a nearly 3-fold increase in cellular glutathione levels and with increased resistance to cell death after treatment with etoposide or SN-38, a derivative of camptothecin, in leukemia 697 cells with wild-type p53. Treatment of Bcl-2-overexpressing 697 cells (697-Bcl-2) with buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, reduced cellular glutathione levels and completely abolished Bcl-2-mediated drug resistance. Morphologic studies revealed that nonapoptotic cell death was induced in 697-Bcl-2 cells after treatment with BSO plus etoposide or SN-38. Activation of caspase-3/7 and cytochrome c release could not be detected in 697-Bcl-2 cells after these drug treatments. Notably, we showed that proteasome-mediated down-regulation of Puma and Noxa proteins occurs in 697-Bcl-2 cells after treatment with BSO plus topoisomerase inhibitor, although there is an increase in the protein levels of p53 in these 697-Bcl-2 cells. In contrast, parental 697 cells underwent typical apoptosis with up-regulation of Puma and Noxa proteins, followed by cytochrome c release and caspase-3/7 activation after treatment with topoisomerase inhibitor in the presence or absence of BSO. Our data suggest that BSO may possess a unique activity to overcome Bcl-2-mediated drug resistance by stimulating the signals that can bypass mitochondrial process in Bcl-2-overexpressing cells. (Cancer Res 2006; 66(11): 5772-80)

We describe a case of Adams-Stokes syncope due to complete atrioventricular block which occurred in a leukemic patient receiving all-trans retinoic acid (ATRA). Remission induction therapy was performed for a 46-year-old Japanese man with acute promyelocytic leukemia using ATRA (45 mg/m2), enocitabine (170 mg/m2, 5 days), and mitoxantrone (4 mg/m2, 3 days). On the 25th day of chemotherapy, syncope suddenly occurred. Electrocardiography revealed a complete atrioventricular block, and a temporary pacemaker was inserted on the following day. The block was persistent and the cardiac rhythm was dependent on the pacemaker. ATRA was discontinued on the 29th day because the arrhythmia was believed to be an adverse reaction to the ATRA regimen. The normal sinus rhythm was restored 15 days thereafter, and the patient eventually reached remission. He subsequently received 4 courses of consolidation therapy without any cardiovascular complications. Although ATRA sometimes induces arrhythmias, to the best of our knowledge this is the first report in the literature of such a critical ATRA-related arrhythmia.

Levels of macrophage colony-stimulating factor (nine of nine patients) and gamma interferon (six of nine patients) in serum were elevated above the range of normal in the acute phase of tsutsugamushi disease. Significant increases in levels of tumor necrosis factor alpha were observed during the convalescent phase in five patients, and they exceeded the levels observed during the acute phase. Hypercytokinemia appeared to be responsible for the emergence of the symptoms of tsutsugamushi disease.

Background Hyperhomocysteinemia is an independent risk factor of coronary artery disease, but some studies have shown that patients with hyperhomocysteinemia are not prone to atherosclerosis. The aim of this study was to test whether homocysteine increases the production of matrix metalloproteinase-2 (MMP-2) and if extracellular additional magnesium and folic acid alters MMP-2 secretion. Methods and Results Gelatin zymography and western blotting were used to investigate the effects of different homocysteine levels (0-5,000 μmol/L) on MMP-2 production, and the effects of different folic acid concentrations (0-10 μmol/L) and magnesium concentrations (0-3.0 mmol/L) on homocysteine-induced MMP-2 in cultured rat vascular smooth muscle cells. Furthermore, the changes in MMP-2 were compared under various treatments for 24 h, 48 h and 72 h. Homocysteine (50-1,000 μmol/L) increased the production of MMP-2 significantly in a dose-dependent manner and at a high level (5,000 μmol/L) reduced the production of MMP-2. Increased production of MMP-2 induced by homocysteine was reduced by additional extracellular folic acid in a dose-dependent manner. Magnesium also reduced the increase of MMP-2 production induced by homocysteine. Production of MMP-2 under various treatments for 72 h increased more than during 24 or 48 h. Conclusions Homocysteine (50-1,000 μmol/L) significantly increased the production of MMP-2 in a dose-dependent manner. Added extracellular folic acid and magnesium decreased the homocysteine-induced MMP-2 secretion. These data suggest a beneficial effect of folic acid and magnesium on the pathogenesis of coronary artery disease. (Circ J 2006; 70: 141 - 146)

L-carnitine (beta-hydroxy-trimethylaminobutyric acid) plays an essential metabolic role that consists of transferring the long chain fatty acids through the mitochondrial barrier, thus allowing their energy-yielding oxidation. GP7 (4-[4''-(2'', 2'', 6'', 6''-tetramethyl-l''-piperidinyloxy) amino] -4'-demethyl-epipodophyllotoxin) is a new spin-labeled derivative of podophyllotoxin semi-synthesized by our university. In this study, we examined the activity of L-carnitine in GP7-induced apoptosis in Burkitt's lymphoma cell line, Raji. GP7 induced time- and dose-dependent apoptotic DNA fragmentation accompanied by caspase-3 activation in Raji cells, and the kinetics of caspase-3 activation induced by GP7 was well correlated with that of apoptotic DNA fragmentation. L-carnitine treatment prevented GP7-induced caspase-3 activation, suppressed caspase-3 cleavage and abrogated GP7-induced apoptotic DNA fragmentation in Raji cells. Our findings suggest that L-carnitine is a potent anti-apoptotic agent to human lymphoma cells and may exert its anti-apoptotic effect via inhibition of caspase-3 activity in GP7-treated Raji cells.

The cytotoxic action of the deoxyadenosine analogue 9-beta-D-arabinofuranosyl-2-fluoroadenine (F-ara-A) depends on the incorporation into DNA after being phosphorylated to F-ara-A triphosphate (F-ara-ATP) by deoxycytidine kinase (dCK). The mechanisms of resistance to F-ara-A were investigated in a newly established variant of L1210 mouse leukemia cells (L1210/F). L1210/F was more than 41-fold more resistant to F-ara-A than the parental cell line and had a 55% lower dCK activity. Interestingly, L1210/F showed a modest level of cross-resistance to deoxycytidine analogues phosphorylated by dCK, for instance, 1-beta-D-arabinofuranosylcytosine (ara-C). The comparative study of F-ara-A and ara-C demonstrated that the difference in the accumulation of their respective triphosphates was minor. In contrast, the incorporation of F-ara-A into DNA was strikingly suppressed compared with that of ara-C. In general, the high natural triphosphate levels interfere with corresponding analogue incorporation into DNA. The deoxyadenosine triphosphate (dATP) and deoxycytidine triphosphate pool sizes in L1210/F cells were increased by 4.9-fold and 1.9-fold, respectively, compared with the parental cells. Treatment with hydroxyurea increased the ratio of F-ara-ATP to dATP 2.1-fold and enhanced the action of F-ara-A in L1210/F. This is the first cell line to show that the profoundly defective incorporation of F-ara-A into DNA during competition with excess dATP confers a high degree of resistance to F-ara-A.

Objective To study the effects of copper and zinc on the production of homocysteine-induced extracellular matrix metalloproteinase-2 in cultured rats vascular smooth muscle cells. Methods We examined the effects of different homocysteine levels (0-1000 micromol/l) on matrix metalloproteinase-2 (MMP-2) production, and the effects of different copper and zinc concentrations (0-300 micromol/l) on homocysteine-induced MMP-2 in cultured rat vascular smooth muscle cells (VSMCs) using gelatin zymography and western blotting. We further compared the changes of MMP-2 under various treatments for 24, 48 and 72 hours. Results Homocysteine (50-1000 micromol/l) increased the production of MMP-2 significantly in a dose-dependent manner. Copper decreased the homocysteine-induced MMP-2 under these conditions in a dose-dependent manner, but zinc did not. Production of MMP-2 under various treatments for 72 h increased more than 24 h and 48 h. Conclusions Extracellular added copper decreased homocysteine-induced MMP-2 secretion.

Several lines of evidence have shown an association between Chlamydia infection and atherosclerosis, but clinical trials of preventive antibiotic (erythromycin) treatment in patients with coronary artery disease have shown conflicting results. Hyperhomocysteinaemia is an independent risk factor of coronary artery disease and causes an intense remodelling of the extracellular matrix in arterial walls, particularly an elastolysis involving metalloproteinases. In the present study we investigated the effects of erythromycin on the production of homocysteineinduced extracellular matrix metalloproteinase-2 (MMP-2) in cultured rat vascular smooth muscle cells (VSMCs).Effects of different concentration of homocysteine (Hcy) (0-5000 micromol/l) on MMP-2 production, and the effects of different erythromycin concentrations (0-10 mmol/l) on homocysteine-induced MMP-2 production in cultured rat VSMCs were studied using gelatin zymography and Western blotting. The changes of MMP-2 under various treatments for 1, 3 and 5 days were also compared.Homocysteine (50-1000 mu mol/l) increased the production of MMP-2 significantly in a dose-dependent manner and reduced the production of MMP-2 at a high level (5000 mu mol/l). Increased production of MMP-2 induced by homocysteine was reduced by extracellularly added erythromycin in a dose-dependent manner.Homocysteine increased the production of MMP-2 significantly in a dose-dependent manner. Extracellularly added erythromycin decreased homocysteine-induced MMP-2 secretion. The findings of the present study suggested that the beneficial effect of erythromycin on vascular disease processes might be due to its inhibitory effect on the Hcyinduced production of MMP-2 in VSMCs.

OBJECTIVES This study sought to clarify the differences of intracellular and extracellular magnesium levels in patients with cardiac syndrome X. METHODS We evaluated the intracellular and extracellular magnesium status of 22 patients with cardiac syndrome X (group A) and 22 age--and gender--matched disorder-free control subjects (group B). Levels of magnesium were determined in serum, urine, erythrocytes, and the 24-h magnesium retention rate was calculated by a magnesium loading test. RESULTS Group A showed a higher 24-h magnesium retention rate (49.8 +/- 1.3% vs. 32.6 +/- 7.5%, p < 0.05) and a lower intracellular concentration of magnesium in erythrocytes than group B (3.7 +/- 1.4 vs. 5.6 +/- 1.3 fg/cell, p < 0.05), demonstrating the presence of magnesium deficiency in group A. There were no significant differences in the serum concentration of magnesium between groups A and B (0.87 +/- 0.23 vs. 0.83 +/- 0.15 mmol/l). CONCLUSIONS This study demonstrated that the intracellular magnesium level decreased in patients with cardiac syndrome X.

To determine the antitumor effect of arsenic trioxide (As2O3) on multidrug-resistant cells, we applied 3 human leukemia cell lines: daunorubicin (DNR)-resistant cell line K562/D1-9, which overexpresses p-glycoprotein (Pgp); DNR and 1-beta-D-arabinofuranosylcytosine (Ara-C) double-resistant cell line HL60/AD, which overexpresses multidrug resistance-associated protein (MRP1); and Bcl-2-transfected pre-B lineage leukemia cell line 697/Bcl-2. Interestingly, K562/D1-9 showed collateral sensitivity. Only HL60/AD showed small cross resistance, but 697/Bcl-2 had no resistance to As2O3. An intracellular content of glutathione (GSH) played a critical role in sensitivity to As2O3. Buthionine-sulfoximine (BSO), which reduces the GSH content, not only increased the As2O3 sensitivity but also conquered the MRP1-related cross resistance in HL60/AD. In conclusion, As2O3 was effective in all 3 cell lines, suggesting that As2O3 may be a promising agent for the treatment of multidrug-resistant leukemia.

OBJECTIVE To study the clinical features, biochemical profiles, echocardiographic and electrocardiographic findings in the elderly and younger with idiopathic dilated cardiomyopathy (IDCM). METHODS We measured biochemical profiles and reviewed the clinical features, echocardiographic and electrocardiographic findings in 40 elderly (group A) and 70 younger (group B) patients with IDCM. RESULTS The aldosterone level in group A was higher than in group B (304.8 +/- 69.1 vs. 213.3 +/- 54.5 pmol/l, P < 0.05). Triiodothyronine (T3) and free T3 in group A were lower than in group B (0.78 +/- 0.21 and 2.87 +/- 0.73 vs. 1.26 +/- 0.33 nmol/l and 3.55 +/- 0.64 pmol/l, all P < 0.05). The incidence of ventricular arrhythmia in group A was lower than in group B (61.3% vs. 92.1 %, P < 0.01). The incidence of hypokalaemia and hypomagnesaemia were higher in group A (51.3% and 27.5%). 28.8% patients in group A were susceptible to digitalis intoxication. Disease duration and mean survival period in group A were longer than in group B (11.0 +/- 4.7 and 6.9 +/- 4.2 vs. 5.2 +/- 2.5 y and 3.4 +/- 2.7 y, all P < 0.05). The main cause of death in group A was congestive heart failure (78.9%) and ventricular arrhythmia (61.9%) in group B. CONCLUSIONS The prognosis in group A was better than in group B. The patients in group A usually showed low T3 syndrome. Ventricular arrhythmia in group A may be due to heart failure, electrolyte imbalance and sympathetic activation.

Cytarabine (ara-C), a major antileukemic agent, is phosphorylated in the cell to cytarabine triphosphate (ara-CTP), which is then partly incorporated into DNA. The drug incorporation into DNA poisons the extending primer against further incorporation of deoxyribonucleotides including dCTP, ultimately inhibiting DNA synthesis. While intracellular ara-CTP concentration has been found to predict clinical outcome, cytotoxicity in vitro is determined primarily by the extent of drug incorporation into DNA. However, clinically appropriate quantitation methods for ara-C at the DNA level have not been available. We developed a sensitive new method for monitoring ara-C incorporated into DNA in vivo. After DNA from leukemic cells was fractionated using the Schmidt-Thannhauser-Schneider method, it was degraded to constituent nucleosides to release ara-C, which was isolated from the nucleosides using HPLC and then measured by radioimmunoassay. Recovery for DNA fractionation, ara-C release by degradation, and ara-C isolation were 92.0+/-6.4%, 90.7+/-9.4%, and 98.5+/-1.4%, respectively. The method was found to determine ara-C incorporation into DNA of ara-C-treated HL 60 cells in vitro with minimal interassay variation. The values determined were compatible with those determined by scintillation counting in parallel experiments using tritiated ara-C. Our method could be used to monitor DNA-incorporated ara-C concentrations during ara-C therapy, together with plasma ara-C and intracellular ara-CTP concentrations. ara-C incorporation into DNA appeared to be associated with intracellular retention of ara-CTP or persistence of plasma ara-C. Thus, the present method is sensitive, accurate, precise, and may permit therapeutic drug monitoring at the DNA level for better individualization of antileukemic regimens.

OBJECTIVES This study aimed to assess whether folic acid supplementation could produce longer-term (6-month) improvements in homocysteine levels and arterial endothelial function in patients with a high risk (3 or more traditional risk factors) of coronary artery disease (CAD) and hyperhomocysteinaemia. METHODS Thirty-one adults with 3 or more traditional risk factors of CAD and hyperhomocysteinaemia were selected, without CAD (the criterion of CAD is that more than one main vessel has an obstruction > or = 50%) by coronary angiography. All subjects were given folic acid (5 mg/day) for 6 months. Fasting plasma homocysteine levels were measured by high-performance liquid chromatography. Plasma folic acid and vitamin B12 levels were measured with immunoassay. Arterial endothelial function was measured as flow-mediated dilation of the brachial artery using high-resolution B-mode ultrasound. RESULTS Folic acid supplementation for 6 months was associated with a significant increase in mean (+/- SD) plasma levels of folic acid (4.6 +/- 1.4 microg/l to 9.1 +/- 2.5 microg/l; P < 0.01) and a significant decline in homocysteine levels (18.3 +/- 3.9 micromol/l to 11.5 +/- 2.8 micromol/l; P < 0.01). Flow-mediated dilation also improved significantly, from 6.8% +/- 2.1% to 8.9% +/- 1.7% (P < 0.01). CONCLUSION These results demonstrate that long-term (6-month) folic acid administration significantly declines homocysteine levels and improves arterial endothelial function and has potential implications for the prevention of atherosclerosis in adults with 3 or more traditional risk factors of CAD and hyperhomocysteinaemia.

Imatinib mesylate (imatinib), a selective inhibitor of BCR-ABL tyrosine kinase, has shown excellent efficacy in patients with chronic myelogenous leukemia (CML) in the chronic phase, however, it does not in those in the accelerated phase or blastic crisis. In patients with CML who have undergone allogeneic stem cell transplantation, imatinib has the capability to induce hematological and even molecular response, and provides a prolonged survival among those in the chronic and accelerated phases. It has been demonstrated that major cytogenic response is a surrogate marker for survival in cases receiving imatinib. It has also been demonstrated that a genome-wide cDNA microarray enables the prediction of sensitivity to imatinib. The acquired resistance in patients who failed to respond to imatinib seemed to be induced by several point mutations in the BCR-ABL gene, which were likely to affect the binding of imatinib with BCR-ABL. Polyclonal cells which harbor distinct mutations in a single patient seemed to be selected in vivo under the selective pressure of imatinib, indicating the rationale of combined treatment with other types of agents. Recently, SPIRIT (STI571 Prospective International Randomized Trials) have been conducted, in which the efficacy of imatinib monotherapy, and imatinib combined with interferon or cytarabine were compared. New agents which inhibit the signaling pathway related to BCR-ABL, such as adaphostin (NSC680410), farnesyltransferase inhibitor SCH66336, MAP kinase inhibitor PD184352, PD98059, U0126, and antibiotic geldanamycin, have shown excellent activity combined with imatinib in vitro.

The cardiac effects of calcium channel blockers (CCBs) related to cardiac remodeling are inconsistent. Matrix metalloproteinases (MMPs) contribute to tissue remodeling. Cardiac fibroblasts play an important role in the regulation of collagen degradation by MMPs. Using gelatin zymography, Western blotting, Griess reagent, and a calcium kit-fluo 3, we investigated the effects of nifedipine, verapamil, diltiazem, and amlodipine on MMP-2 expression and further elucidate the mechanisms in cultured rat cardiac fibroblasts. Nifedipine increased and amlodipine decreased the expression of MMP-2; however, neither verapamil nor diltiazem altered MMP-2 expression. Nifedipine also increased nitrite production, and this increase was blunted by a nitric oxide (NO) synthases inhibitor (L-NAME). Nifedipine-induced MMP-2 expression was also blunted by L-NAME. An NO donor (sodium nitroprusside) induced MMP-2 expression. Data indicated that nifedipine might increase MMP-2 expression through a possible NO-dependent pathway. Amlodipine had no influence on nitrite production. The amlodipine-induced decrease of MMP-2 expression was abolished by two protein tyrosine kinase inhibitors, genistein and herbimycin A, indicating that amlodipine might decrease MMP-2 expression through a possible protein tyrosine kinase pathway. None of the four CCBs could alter the fluoscence intensity of fluo 3, indicating that the effects of CCBs on MMP-2 expression were independent of the variation in intracellular C2+ concentration. Our findings revealed that different CCBs exerted different effects on MMP-2 expression in cardiac fibroblasts.

To study the acute results and long-term clinical course after percutaneous transluminal septal myocardial ablation (PTSMA) in symptomatic patients with hypertrophic obstructive cardiomyopathy (HOCM).In 18 patients (seven women, 11 men; average age 53+/-15 years) with symptomatic and medically refractory HOCM, 1.3+/-0.4 septal branches were occluded with an injection of 3.6+/-1.2 mL of 95% alcohol to ablate the hypertrophied interventricular septum. After three years, noninvasive follow-up results of clinical course, echocardiographic and electrocardiographic findings were determined.The invasively measured left ventricular outflow tract (LVOT) gradients were reduced in all patients, with a mean decrease from 79+/-21 mmHg to 11+/-8 mmHg at rest (P0.01) and from 136+/-41 mmHg to 49+/-21 mmHg after extrasystole. All patients had angina pectoris for 8 h to 24 h. Eleven patients (61.1%) developed a trifascicular block for 3 min to four days requiring temporary (n=10 [56%]) or permanent dual chamber pacemaker implantation (n=1 [6%]). All patients were discharged after 5.9+/-2.3 days. Clinical, electrocardiographic and echocardiographic follow-up was achieved in all patients after three years (3.1+/-0.5 years). No cardiac complications occurred. Thirteen patients (72%) showed clinical improvement, with a New York Heart Association functional class of 1.5+/-0.8. A further reduction in LVOT gradient was shown in eight patients (44%).The LVOT gradient was greatly reduced in patients with HOCM undergoing a PTSMA procedure and their symptoms were greatly improved without cardiac complications during three-year follow-up. Possible complications include different degrees of heart block, such as trifascicular blocks, requiring temporary pacemaker implantation. PTSMA is a promising nonsurgical method for the treatment of symptomatic patients with HOCM. Clinical long-term follow-up of a larger series of patients is required to determine the therapeutic significance conclusively.

The effects of fuel octane have been assessed on the efficiency and emissions of a high compression ratio (e=13) spark ignition direct injection (SIDI) engine. Under low load stratified operation (1200 rpm, ∼20% load), a low octane fuel (RON=84, comprised of toluene, iso-octane, and n-heptane) yielded higher brake thermal efficiency and significantly lower hyd carbon emissions than a base gasoline (RON=91). The indicator diagram for the low octane fuel showed evidence for two stage heat release, suggesting the presence of spark induced compression ignition (SICI). These results suggest that higher efficiency under low load stratified conditions can be obtained with lower octane fuels that undergo SICI combustion. The effect of fuel octane under high load was assessed at WOT with a high RON model fuel (RON=103, comprised of toluene, iso-octane, and n-heptane). This high octane fuel exhibited a torque benefit compared to pure iso-octane (RON=100) and premium gasoline (RON=99) that is significantly greater than expected based on RON alone. The results suggest that a high RON fuel, in particular one that is high in aromatics, yields significant torque benefits under high load.

Complete elimination of the plasma cell dyscrasia is a rational therapeutic goal, as intercepting supply of precursor protein is a necessary condition for a major regression of amyloid deposits. High-dose melphalan with autologous stem cell transplantation has shown the ability to induce complete hematological response (HR) along with recovery of organ dysfunction. However, the rate of HR with this treatment rarely exceeds 40%. We describe here the first known case of successful reduced intensity allogeneic stem cell transplantation (RIST) for a patient with primary amyloidosis complicated with nephrotic syndrome but without cardiac disease, who had obtained only partial HR by high-dose melphalan with autologous stem cell transplantation. RIST may be feasible and be capable of achieving complete HR along with recovery from nephrotic syndrome with acceptable toxicity.

Two antibiotics recommended by the guideline of Infectious Diseases Society of America (IDSA) were selected for treatment of febrile neutropenia, and these paired antibiotics were changed periodically three times. The clinical efficacy of each antibiotic was retrospectively evaluated at the end of the final period. There was no significant difference about efficacy rate between two kinds of antibiotics in the same sequential period. However, the efficacy rate has been rising and febrile duration has been shortening by degrees. Only a few drug resistant bacteria were recognized by the surveillance culture during antibiotic cycling. Recently, antibiotic cycling therapy has attracted attention especially in the ICU. However, a clinical study of treatment for febrile neutropenia has not been reported. Our trial suggests that cycling therapy may be useful for febrile neutropenia. However, Some deviation in the patients characteristics of each period may affect the result. It seems that further examination is necessary about usefullness of the cycling therapy for febrile neutropenia.

Many traditional independent risk factors such as diabetes mellitus, hypertension, hypercholesterolaemia, smoking, male sex, old age, etc., contribute to the development of coronary artery disease (CAD). Hyperhomocysteinaemia is an independent risk factor of CAD but the role of plasma homocysteine (Hcy) in high risk patients (or = 3 risk factors) is not known. We investigated the role of plasma Hcy, folic acid, vitamin B12 in patients with high risk (or = 3 risk factors) of CAD and effects of supplementation of folic acid in the patients with hyperhomocysteinaemia.The plasma Hcy levels in 152 patients withor = 3 risk factors of CAD and 136 patients with 1-2 risk factors and 48 individuals with no risk factors were measured using high performance liquid chromatography (HPLC) with fluorescence detection. Plasma folic acid and vitamin B12 levels were also measured in these patients with immunoassays. The patients with hyperhomocysteinaemia were treated with 5 mg of folic acid for 8 wk, and plasma levels of Hcy were measured after treatment.The plasma Hcy level was significantly higher in the patients withor = 3 risk factors of CAD than in those with 1-2 risk factors and controls. The plasma levels of folic acid and vitamin B12 were significantly lower in the patients withor = 3 risk factors of CAD compared to those with 1-2 risk factors and controls. The Hcy levels in the patients withor = 3 risk factors of CAD significantly reduced by 33.5 per cent after 8 wk folic acid administration.Plasma Hcy level was elevated significantly in patients withor = 3 risk factors of CAD. Hyperhomocysteinaemia appears to play an important role in the pathogenesis of CAD. Folic acid supplementation may be useful in reducing plasma Hcy level in high risk patients with hyperhomocysteinaemia.

Instead of the original intermediate dose (0.5 g/m2), a modified intermediate-dose 1-beta-D-arabinofuranosylcytosine (ara-C) therapy (1 g/m2, 1-h intravenous infusion) was pharmacologically studied in 11 leukemic patients.The concentrations of ara-C and its inactive metabolite, 1-beta-D-arabinofuranosyluracil (ara-U) in the plasma, urine and cerebrospinal fluid were determined using high performance liquid chromatography.The plasma ara-C reached a peak (61.2 +/- 52.7 microM) that far surpassed the saturating level for intracellular activation of the drug. The plasma ara-U reached its peak (139.8 +/- 40.0 microM) and was maintained with a half-life of 277 +/- 76 min. About 60% of the administered drug was recovered, mainly as ara-U in urine within 12 h. In contrast to the original intermediate dose, the therapeutic ara-C levels (above 0.4 microM) persisted in the central nervous system. The therapy salvaged one relapsed leukemia patient with central nervous system involvement.Modified intermediate-dose ara-C provides a sufficient plasma ara-C level with a concomitant therapeutic concentration in the cerebrospinal fluid.

Previous studies including ours have demonstrated that DNA repair is one of the important targets of fludarabine. The aim of this study is to clarify a mechanistic interaction of carboplatin and F-ara-A, from the perspective of F-ara-A-mediated inhibition of DNA repair initiated by carboplatin. Using human quiescent lymphocytes, we focused on DNA repair, since these cells provide a model of dormant cells. To evaluate the carboplatin-induced DNA incision and its repair, we used the alkaline comet assay. When lymphocytes were incubated with carboplatin, a dose-dependent increase in the tail-moment was observed. Then, tail-moment decreased in proportion to the incubation period in fresh media and recovered to the control level at 4 h. DNA rejoining was completely inhibited by F-ara-A at 10 microM through 0 to 6 h after washing out of these drugs and this F-ara-A-induced inhibition was concentration-dependent. Cellular damage after drug exposure was evaluated with the induction of apoptosis as well as cytotoxic effect. Exposure to carboplatin alone did not induce any apparent cellular damage in quiescent lymphocytes. In contrast, a more than additive induction of apoptosis as well as an enhancement of cytotoxic action was observed in cells treated with a combination of carboplatin and F-ara-A. In the CEM cell line, there was no enhancement of the cytotoxic action of these drugs, despite the clear demonstration of an inhibitory effect on DNA repair. These results indicate that chemotherapy with carboplatin opened a new target for F-ara-A by initiating DNA repair in quiescent cells.

Survivin, a new member of the inhibitor of apoptosis protein (IAP) family, has been reported to be expressed in many cancers but not in differentiated normal tissues. In the present study, we describe the identification of a novel alternatively spliced survivin transcript, designated as survivin-3B. It comprises 5 exons including novel exon 3B derived from a 165-bp long portion of intron 3. Acquisition of a new in-frame TGA stop codon within the novel exon 3B results in an open reading frame (ORF) of 363 nucleotides, predicting a truncated 120 amino acid protein. Expression of survivin-3B was detected in human colon and gastric adenocarcinoma cell lines as well as mononuclear cells prepared from patients with myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). Survivin-3B contains a single baculovirus IAP repeat (BIR), which is critical for apoptosis inhibition. However, it lacks a carboxyl-terminal coiled-coil domain, suggesting that survivin-3B may not be associated with G2/M phase. These data indicate that the function of survivin-3B may be different from that of regular survivin.

The aim of this study was to study the relationship between plasma homocysteine (Hcy), folic acid, vitamin B12 and early coronary artery disease (early-CAD) and high coronary risk factors. The plasma Hcy levels of 58 cases with early-CAD and 31 subjects without CAD were measured using high-performance liquid chromatography (HPLC) with fluorescence detection. Plasma folic acid and Vitamin B12 levels were measured with radioassay method. The plasma Hcy level was significantly higher in the early-CAD patients than in the controls [(13.7 +/- 5.1) micromol/L vs (10.3 +/- 5.7) micromol/L]. The plasma folic acid and Vitamin B12 levels were significantly lower in the early-CAD patients than in the controls. The plasma Hcy level was higher in patients with more than 3 risk factors of CAD than in patients with 1 or 2 risk factors and in the controls [(17.3 +/- 5.1) micromol/ L vs (12.9 +/- 4.8) micromol/L, (7.8 +/- 2.5) micromol/L]. Other than gender, all classical coronary risk factors were related to the elevated Hcy level. Hcy levels were elevated in patients with early-CAD and with high risk factors. Hyperhomocysteinemia plays an important role in the pathogenesis of CAD.

GP7 (4-[4"-(2",2",6",6"-tetramethyl-l"-piperidinyloxy)amino]-4'-demethyl epipodophyllotoxin), a new spin-labeled derivative of podophyllotoxin, is a promising anticancer drug of podophyllotoxin class. The primary effect of GP7 is the anticancer activity on transplanted mouse tumors and cultured tumor cells. However, its molecular mechanism of action is still obscure. In this study, we investigated the activity of GP7 to induce apoptosis in human leukemia HL-60 and Jurkat cells. Apoptosis was determined by detection of DNA fragmentation in agarose gel electrophoresis. GP7 induced apoptotic DNA fragmentation of HL-60 and Jurkat cells in time- and dose-dependent manner. We further investigated the activity of caspase-3 in GP7-induced apoptotic DNA fragmentation of HL-60 and Jurkat cells. GP7 also induced time- and dose-dependent caspase-3 activation in both cell lines, and the kinetics of caspase-3 activation induced by GP7 was well correlated with that of apoptotic DNA fragmentation. To determine the role of caspase-3 in GP7-induced apoptotic DNA fragmentation, we examined the effect of specific caspase-3 inhibitor, Ac-DEVD-CHO, on GP7-induced apoptotic DNA fragmentation. Ac-DEVD-CHO prevented GP7-induced caspase-3 activation in both HL-60 and Jurkat cells, however, it only inhibited GP7-induced apoptotic internucleosomal DNA fragmentation in HL-60 cells. We then employed L-carnitine to investigate the role of caspase-3 in GP7-induced apoptotic DNA fragmentation. L-carnitine treatment prevented GP7-induced caspase-3 activation in both cell lines in a dose-dependent manner. Similar to Ac-DEVD-CHO, L-carnitine only inhibited GP7-induced apoptotic internucleosomal DNA fragmentation in HL-60 cells. These findings suggest that GP7 exerts an anti-leukemic effect by both caspase-3-dependent and -independent apoptotic signaling pathways.

A simple, isocratic HPLC method was newly developed for quantitating intracellular fludarabine triphosphate (F-ara-ATP). Samples (500 microl) were injected onto an anion-exchange column and eluted isocratically with phosphate-acetonitrile buffer (flow rate: 0.7 ml/min) at an ambient temperature. F-ara-ATP was quantitated according to its peak area at the absorbance of 261 nm. The standard curve was linear with minimal within-day and inter-day variability. The low and high quantification limits were 50 pmol and 20 nmol, respectively. The method was capable of measuring F-ara-ATP generated in cultured leukemic cells in vitro. Thus, our method will be useful because of its sensitivity and simplicity as well as applicability to biological materials.

Spark ignition direct injection (SIDI) engines have the potential to realize significant thermal efficiency improvements compared to conventional port fuel injection engines. The effects of fuel properties on efficiency and emissions have been investigated in a prototype of an Avensis Wagon equipped with a 2.0 liter, 4 cylinder spark ignition, direct injection (SIDI) engine designed to meet US 2000 emission standards. The vehicle employed a close coupled three-way catalyst and a NOx storage and reduction catalyst. Seven matrix fuels were blended to the same RON with varying levels of aromatics, olefins, ethanol, and volatility. Relative thermal efficiency, fuel economy, and tailpipe emissions were measured for the matrix fuels and a base fuel under the FTP LA4 driving cycle. The engine was operated in a lean burn mode in light load condition for approximately half of the driving cycle. Fixed speed/load engine bench tests were also carried out for three of the matrix fuels to complement the vehicle tests. In addition, laminar burning velocity measurements of all seven matrix fuels were made in a constant volume combustion vessel. The vehicle tests showed a 2% spread in relative thermal efficiency, with increased efficiency correlating with higher olefins and lower aromatics. In the bench engine tests, a 2 % increase in peak torque was observed, with the fuel having the highest olefins plus aromatics content yielding the highest torque. The influence of the primary fuel properties autoignition resistance and burn rate (laminar burning velocity) on these results is discussed. Lower aromatic levels directionally correlated with decreased NOx emissions, and lower driveability index (DI) directionally correlated with decreased non-methane hydrocarbon emissions.

The precise correlation between magnesium and cardiac disease remains to be established. Matrix metalloproteinases (MMPs) are important in cardiac disease such as heart failure. Cardiac fibroblasts are the most abundant cell type in the heart and play an important role in the regulation of collagen degradation by MMPs. To assess the association between magnesium and MMPs, we examined the effects of different extracellular magnesium concentrations (0-3.0 mmol/L) on MMP-2 production in cultured rat cardiac fibroblasts. Using gelatin zymography and western blotting, we found that magnesium reduced MMP-2 production dose-dependently, and this effect was inhibited by the tyrosine kinase inhibitors, genistein or herbimycin A. The results of this study indicated that the beneficial effect of magnesium supplementation on the cardiac disease may be due, at least in part, to the inhibitory effect of magnesium on production of MMPs in cardiac fibroblasts, which appears to be mediated by a protein tyrosine phosphorylation related signal transduction pathway.