Effect of basal insulin therapy on vascular endothelial function and adipokine profiles in people with Type 2 diabetes.

AIM: To compare the effects of the basal insulin analogues glargine and detemir on endothelial function and adipocytokine levels in people with Type 2 diabetes. METHODS: We studied 32 people with Type 2 diabetes whose blood glucose control was unsatisfactory while receiving only oral hypoglycaemic drugs. Participants were randomized to either insulin glargine or detemir for 24 weeks and then crossed over to the other treatment without a washout period. Flow-mediated vasodilatation, adipocytokine levels (plasminogen activator inhibitor-1 and leptin/adiponectin ratio), and fasting ghrelin levels were monitored. RESULTS: HbA1c levels were significantly decreased by both basal insulin therapies. Body weight was significantly increased by glargine but not by detemir. The proportion of flow-mediated vasodilatation was significantly increased by detemir but not glargine (glargine: from 5.17 ± 0.69 to 5.94 ± 0.83%; detemir: from 4.89 ± 0.78 to 7.92 ± 0.69%). Plasminogen activator inhibitor-1 level was significantly decreased by only detemir (glargine: from 16.4 ± 1.8 to 17.3 ± 2.1; detemir: from 19.2 ± 2.8 to 16.0 ± 1.6 ng/ml). The leptin/adiponectin ratio was significantly increased only by glargine. Acyl ghrelin level was significantly decreased by glargine but not detemir. CONCLUSIONS: These results suggest that the effect on endothelial function and adipocytokine profiles may differ between glargine and detemir in people with diabetes (Trial registration ID: UMIN000004973).

Impact of B-type natriuretic peptide (BNP) on development of atrial fibrillation in people with Type 2 diabetes.

AIMS: To examine if a simple biomarker can identify people with diabetes who are at high risk of atrial fibrillation. METHODS: A retrospective cohort study was conducted at a single centre in people with Type 2 diabetes referred to our department between January 2000 and December 2007. In 517 consecutive people without any history, signs or symptoms of atrial fibrillation at baseline, the association between baseline B-type natriuretic peptide level and future atrial fibrillation incidence was examined, with adjustments for other potentially confounding factors. RESULTS: A total of 28 people were diagnosed with new-onset atrial fibrillation during a median 6-year follow-up. When people were categorized into three groups according to B-type natriuretic peptide clinical thresholds (20 and 100 pg/ml), hazard ratios for the development of atrial fibrillation in the middle and highest B-type natriuretic peptide groups were 2.8 and 9.4, respectively, compared with the lowest B-type natriuretic peptide group. Time-dependent receiver-operating curve analysis identified a threshold for B-type natriuretic peptide to detect atrial fibrillation development of 52.8 pg/ml (sensitivity 75.2%, specificity 68.8%). The B-type natriuretic peptide predictive value was independent of and similar to that of left atrial size and ventricular dimension. CONCLUSION: In people with Type 2 diabetes, high baseline B-type natriuretic peptide levels were significantly associated with future atrial fibrillation development.

Induced pluripotent stem cells generated from diabetic patients with mitochondrial DNA A3243G mutation.

AIMS/HYPOTHESIS: The aim of this study was to generate induced pluripotent stem (iPS) cells from patients with mitochondrial DNA (mtDNA) mutation. METHODS: Skin biopsies were obtained from two diabetic patients with mtDNA A3243G mutation. The fibroblasts thus obtained were infected with retroviruses encoding OCT4 (also known as POU5F1), SOX2, c-MYC (also known as MYC) and KLF4. The stem cell characteristics were investigated and the mtDNA mutation frequencies evaluated by Invader assay. RESULTS: From the two diabetic patients we isolated four and ten putative mitochondrial disease-specific iPS (Mt-iPS) clones, respectively. Mt-iPS cells were cytogenetically normal and positive for alkaline phosphatase activity, with the pluripotent stem cell markers being detectable by immunocytochemistry. The cytosine guanine dinucleotide islands in the promoter regions of OCT4 and NANOG were highly unmethylated, indicating epigenetic reprogramming to pluripotency. Mt-iPS clones were able to differentiate into derivatives of all three germ layers in vitro and in vivo. The Mt-iPS cells exhibited a bimodal degree of mutation heteroplasmy. The mutation frequencies decreased to an undetectable level in six of 14 clones, while the others showed several-fold increases in mutation frequencies (51-87%) compared with those in the original fibroblasts (18-24%). During serial cell culture passage and after differentiation, no recurrence of the mutation or no significant changes in the levels of heteroplasmy were seen. CONCLUSIONS/INTERPRETATION: iPS cells were successfully generated from patients with the mtDNA A3243G mutation. Mutation-rich, stable Mt-iPS cells may be a suitable source of cells for human mitochondrial disease modelling in vitro. Mutation-free iPS cells could provide an unlimited, disease-free supply of cells for autologous transplantation therapy.

Decreased circulating CD34+ cells are associated with progression of diabetic nephropathy.

AIMS: Circulating progenitor cells such as CD34+ cells play a key role in maintenance of vascular endothelial function and neovascularization, and a decrease in the number of CD34+ cells is associated with cardiovascular disease. However, the contribution of circulating progenitor cells to microvascular disease, such as diabetic nephropathy, is unclear. This study was therefore designed to clarify the association between diabetic nephropathy and circulating CD34+ cells. METHODS: We measured circulating CD34+ cell numbers in 85 Type 2 diabetic patients aged 40-70 years with normo- and microalbuminuria and determined the association with urinary albumin excretion rate (UAER). RESULTS: The number of circulating CD34+ cells significantly correlated with log UAER (r = -0.289, P = 0.008). Furthermore, in patients with low numbers of CD34+ cells (0.68 > cells/microl, lowest quartile of CD34+ cell number) UAER increased significantly after 12 months compared with baseline [from 34.3 +/- 7.0 to 53.6 +/- 10.8 mg/g creatinine (gCr), P < 0.05], whereas in patients with a high number of CD34+ cells (1.0 < cells/microl, highest quartile of CD34+ cell number) UAER did not change (from 16.7 +/- 4.8 to 20.1 +/- 3.0 mg/gCr). CONCLUSIONS: These results suggest that a decreased number of circulating CD34+ cells is involved in the progression of diabetic nephropathy and may be a predictor of the disease.

SIRT2, a tubulin deacetylase, acts to block the entry to chromosome condensation in response to mitotic stress.

We previously identified SIRT2, an nicotinamide adenine dinucleotide (NAD)-dependent tubulin deacetylase, as a protein downregulated in gliomas and glioma cell lines, which are characterized by aneuploidy. Other studies reported SIRT2 to be involved in mitotic progression in the normal cell cycle. We herein investigated whether SIRT2 functions in the mitotic checkpoint in response to mitotic stress caused by microtubule poisons. By monitoring chromosome condensation, the exogenously expressed SIRT2 was found to block the entry to chromosome condensation and subsequent hyperploid cell formation in glioma cell lines with a persistence of the cyclin B/cdc2 activity in response to mitotic stress. SIRT2 is thus a novel mitotic checkpoint protein that functions in the early metaphase to prevent chromosomal instability (CIN), characteristics previously reported for the CHFR protein. We further found that histone deacetylation, but not the aberrant DNA methylation of SIRT2 5'untranslated region is involved in the downregulation of SIRT2. Although SIRT2 is normally exclusively located in the cytoplasm, the rapid accumulation of SIRT2 in the nucleus was observed after treatment with a nuclear export inhibitor, leptomycin B and ionizing radiation in normal human fibroblasts, suggesting that nucleo-cytoplasmic shuttling regulates the SIRT2 function. Collectively, our results suggest that the further study of SIRT2 may thus provide new insights into the relationships among CIN, epigenetic regulation and tumorigenesis.

Tumour budding at invasive margins and outcome in colorectal cancer.

OBJECTIVE: Tumour budding, defined as small clusters of undifferentiated cancer cells at invasive margins, has been shown to reflect biologic aggressiveness of colorectal cancers. We therefore examined the prognostic significance of tumour budding in patients with colorectal carcinoma, particularly focusing on comparisons with other clinicopathological findings. METHOD: Tumour budding was investigated in surgically resected specimens from 159 patients with colorectal carcinoma. With haematoxylin and eosin stained slides containing the entire invasive margin, the degree of tumour budding was classified into three grades: mild, <1/3 of the entire invasive margin; moderate, 1/3-2/3; marked, >2/3. RESULTS: Mild tumour budding was found in 54 (34%) cases, moderate in 59 (37%) cases and marked in 46 (29%) cases. The degree of budding was linked with poor tumour differentiation, lymph node metastasis and advanced TNM stage (P < 0.001). In univariate analysis, patients with marked tumour budding [5-year cancer-related survival (CRS)/recurrence-free survival (RFS), 39%/53%] had significantly worse survival [CRS, hazard ratio (HR), 4.561; 95% confidence interval (CI), 2.265-9.184; P < 0.001; RFS, HR, 3.240; 95% CI, 1.430-7.342; P = 0.005] than those with mild (5-year CRS/RFS, 80%/82%) or moderate (63%/66%) budding. In the Cox regression model, marked tumour budding (HR, 3.137; 95% CI, 1.517-6.487; P = 0.002) and advanced tumour stage (stage III, HR, 3.226; 95% CI, 1.475-7.053; P = 0.003; stage IV, HR, 24.443; 95% CI, 10.843-55.100; P < 0.001) proved to be an independent predictor of short CRS. CONCLUSION: Tumour budding is a practical and significant histological index for identification of high malignant potential and poor outcome in patients with colorectal carcinoma.

Rapid transcriptional activation and early mRNA turnover of brain natriuretic peptide in cardiocyte hypertrophy. Evidence for brain natriuretic peptide as an "emergency" cardiac hormone against ventricular overload.

We previously demonstrated that brain natriuretic peptide (BNP) is a cardiac hormone mainly produced in the ventricle, while the major production site of atrial natriuretic peptide (ANP) is the atrium. To assess the pathophysiological role of BNP in ventricular overload, we have examined the gene expression of BNP, In comparison with that of ANP, in a model of cardiac hypertrophy using cultured neonatal rat ventricular cardiocytes. During cardiocyte hypertrophy evoked by endothelin-1, Phenylephrine, or PMA, the steady state level of BNP mRNA increased as rapidly as the "immediate-early" induction of the c-fos gene expression, and reached a maximal level within 1 h. Actinomycin D, a transcriptional inhibitor, completely diminished the response, while the translational blocked with cycloheximide did not inhibit it. In contrast, ANP mRNA began to increase 3 h after the stimulation, and accumulated during cardiocyte hypertrophy. The BNP secretion from ventricular cardiocytes was also stimulated, more rapidly than the ANP secretion. Furthermore, the turnover of BNP mRNA was significantly faster than that of ANP mRNA, being consistent with the existence of AUUUA motif in the 3'-untranslated region of BNP mRNA. These results demonstrate that the gene expression of BNP is distinctly regulated from that of ANP at transcriptional and posttranscriptional levels, and indicate that the characteristics of the BNP gene expression are suitable for its possible role as an " emergency" cardiac hormone against ventricular overload.

Molecular cloning of hamster brain and atrial natriuretic peptide cDNAs. Cardiomyopathic hamsters are useful models for brain and atrial natriuretic peptides.

Brain and atrial natriuretic peptides (BNP and ANP) are cardiac hormones with diuretic, natriuretic, and vasodilatory activities. Cardiomyopathic hamsters are widely used animal models of heart failure. Due to the structural divergence of BNP among species, examination on pathophysiological roles of BNP using cardiomyopathic hamsters is so far impossible. We therefore isolated hamster BNP and ANP cDNAs, and investigated synthesis and secretion of these peptides in normal and cardiomyopathic hamsters. The COOH-terminal 32-residue peptide of cloned hamster preproBNP with 122 amino acids, preceded by a single arginine residue, supposedly represents hamster BNP showing < 50% homology to rat BNP. Alpha-hamster ANP, 28-residue peptide, is identical to alpha-rat ANP. In hamsters, BNP and ANP occur mainly in the ventricle and the atrium, respectively. The 32-wk-old hypertrophic cardiomyopathic BIO14.6 strain exhibited ventricular hypertrophy. The 32-wk-old dilated cardiomyopathic BIO53.58 strain remained at the stage without apparent heart failure. In BIO14.6 and BIO53.58 strains at this age, ventricular BNP and ANP gene expressions are augmented, and the plasma BNP concentration is elevated to 136 and 108 fmol/ml, respectively, three times greater than the elevated plasma ANP concentration, which well mimics changes of the plasma BNP and ANP concentrations in human heart failure. Cardiomyopathic hamsters, therefore, are useful models to investigate the implication of BNP in human cardiovascular diseases.

Blockade of the natriuretic peptide receptor guanylyl cyclase-A inhibits NF-kappaB activation and alleviates myocardial ischemia/reperfusion injury.

Acute myocardial infarction (AMI) remains the leading cause of death in developed countries. Although reperfusion of coronary arteries reduces mortality, it is associated with tissue injury. Endothelial P-selectin-mediated infiltration of neutrophils plays a key role in reperfusion injury. However, the mechanism of the P-selectin induction is not known. Here we show that infarct size after ischemia/reperfusion was significantly smaller in mice lacking guanylyl cyclase-A (GC-A), a natriuretic peptide receptor. The decrease was accompanied by decreases in neutrophil infiltration in coronary endothelial P-selectin expression. Pretreatment with HS-142-1, a GC-A antagonist, also decreased infarct size and P-selectin induction in wild-type mice. In cultured endothelial cells, activation of GC-A augmented H2O2-induced P-selectin expression. Furthermore, ischemia/reperfusion-induced activation of NF-kappaB, a transcription factor that is known to promote P-selectin expression, is suppressed in GC-A-deficient mice. These results suggest that inhibition of GC-A alleviates ischemia/reperfusion injury through suppression of NF-kappaB-mediated P-selectin induction. This novel, GC-A-mediated mechanism of ischemia/reperfusion injury may provide the basis for applying GC-A blockade in the clinical treatment of reperfusion injury.

The neuron-restrictive silencer element-neuron-restrictive silencer factor system regulates basal and endothelin 1-inducible atrial natriuretic peptide gene expression in ventricular myocytes.

Induction of the atrial natriuretic peptide (ANP) gene is a common feature of ventricular hypertrophy. A number of cis-acting enhancer elements for several transcriptional activators have been shown to play central roles in the regulation of ANP gene expression, but much less is known about contributions made by transcriptional repressors. The neuron-restrictive silencer element (NRSE), also known as repressor element 1, mediates repression of neuronal gene expression in nonneuronal cells. We found that NRSE, which is located in the 3' untranslated region of the ANP gene, mediated repression of ANP promoter activity in ventricular myocytes and was also involved in the endothelin 1-induced increase in ANP gene transcription. The repression was conferred by a repressor protein, neuron-restrictive silencer factor (NRSF). NRSF associated with the transcriptional corepressor mSin3 and formed a complex with histone deacetylase (HDAC) in ventricular myocytes. Trichostatin A (TSA), a specific HDAC inhibitor, relieved NRSE-mediated repression of ANP promoter activity, and chromatin immunoprecipitation assays revealed the involvement of histone deacetylation in NRSE-mediated repression of ANP gene expression. Furthermore, in myocytes infected with recombinant adenovirus expressing a dominant-negative form of NRSF, the basal level of endogenous ANP gene expression was increased and a TSA-induced increase in ANP gene expression was apparently attenuated, compared with those in myocytes infected with control adenovirus. Our findings show that an NRSE-NRSF system plays a key role in the regulation of ANP gene expression by HDAC in ventricular myocytes and provide a new insight into the role of the NRSE-NRSF system outside the nervous system.

Induction of JAB/SOCS-1/SSI-1 and CIS3/SOCS-3/SSI-3 is involved in gp130 resistance in cardiovascular system in rat treated with cardiotrophin-1 in vivo.

CIS (cytokine-inducible SH2 protein), SOCS (suppressor of cytokine signaling), or SSI (signal transducers and activators of transcription [STAT]-induced STAT inhibitor) proteins are a family of cytokine-inducible negative regulators of cytokine signaling via Janus kinase (JAK)-STAT pathways. Given the evidence that the JAK-STAT pathway plays a critical role in the cardiovascular system, the primary objective of this study was to assess the effects of the CIS family on JAK-STAT signaling in the cardiovascular system in rats treated with cardiotrophin-1 (CT-1), an interleukin-6 family of cytokines. Intravenous injection of 20 microgram/kg body weight of CT-1 induced a transient, marked increase in STAT3 activation in various tissues, including heart and lung, and subsequent upregulation of 2 members of the CIS family, JAK-binding protein (JAB)/SOCS-1/SSI-1 and CIS3/SOCS-3/SSI-3, in the same tissues. It was also observed that CIS3 was directly associated with JAK2 in vivo. Pretreatment with the same dose of CT-1 60 minutes before significantly attenuated the STAT3 activation induced by a second injection of CT-1. We previously reported that intravenous injection of CT-1 results in the nitric oxide (NO)-dependent hypotension accompanied by the induction of inducible NO synthase mRNA. In rats pretreated with CT-1, the induction of inducible NO synthase mRNA or hypotension by subsequent CT-1 injection was not observed. Forced expression of JAB or CIS3, but not other CISs, directly blocked CT-1-induced STAT3 activation in 293 cells. These results suggest that JAB and CIS3 serve as endogenous inhibitors of CT-1-mediated JAK-STAT signaling in the cardiovascular system in vivo.

Germline deletion and a somatic mutation of the PRKAR1A gene in a Carney complex-related pituitary adenoma.

OBJECTIVE: The objective was to assess involvement of loss of the PRKAR1A gene encoding a type 1α regulatory subunit of cAMP-dependent protein kinase A located on 17q24 in a Carney complex (CNC)-related pituitary adenoma. DESIGN: We investigated aberrations of the PRKAR1A gene in a CNC patient with a GH-producing pituitary adenoma, whose family has three other members with probable CNC. METHODS: A gene mutation was identified by a standard DNA sequencing method based on PCR. DNA copy number was measured to evaluate allelic loss on 17q24 by quantitative PCR. The breakpoints of deletion were determined by cloning a rearranged region in the deleted allele. RESULTS: A PRKAR1A mutation of c.751_758del8 (p.S251LfsX16) was found in genomic DNA obtained from a pituitary adenoma, but not leukocytes from the patient. Reduced DNA copy number at loci including the PRKAR1A gene on 17q24 was detected in both the tumor and leukocytes, suggesting a deletion at the loci at the germline level. The deletion size was determined to be ∼ 0.5 Mb and this large deletion was also found in two other family members. CONCLUSION: This is the first case showing a CNC-related pituitary adenoma with the combination of somatic mutation and a large inherited deletion of the PRKAR1A gene. Biallelic inactivation of PRKAR1A appears to be necessary for the development of CNC-related pituitary adenoma.

Genetic models reveal that brain natriuretic peptide can signal through different tissue-specific receptor-mediated pathways.

Brain natriuretic peptide (BNP), a hormone produced primarily by the cardiac ventricle, is thought to be involved in a variety of homeostatic processes through its cognate receptor, guanylyl cyclase A (GC-A). We previously created transgenic mice overexpressing BNP under the control of the liver-specific human serum amyloid P component promoter (BNP-transgenic mice) and demonstrated that they exhibit reduced blood pressure and cardiac weight accompanied by an elevation of plasma cGMP concentrations and marked skeletal overgrowth through the activation of endochondral ossification. To address whether BNP exerts its biological effects solely through GC-A, we produced BNP-transgenic mice lacking GC-A (BNP-Tg/GC-A-/- mice) and examined their cardiovascular and skeletal phenotypes. The GC-A-/- mice are hypertensive with cardiac hypertrophyrelative to wild-type littermates, which is not alleviated by overexpression of BNP in BNP-Tg/GC-A-/- mice. The BNP-Tg/GC-A-/- mice, however, continue to exhibit marked longitudinal growth of vertebrae and long bones comparably to BNP-Tg mice. This study provides genetic evidence that BNP reduces blood pressure and cardiac weight through GC-A, whereas it dramatically alters endochondral ossification in the absence of this receptor. Therefore, the BNP-Tg/GC-A-/- mice provide the first experimental model demonstrating that this natriuretic peptide can signal in a tissue-specific manner through a receptor other than GC-A.

Identification and endothelin-induced activation of multiple extracellular signal-regulated kinases in aortic smooth muscle cells.

In order to explore intracellular signaling pathways of the mitogenic action of endothelin (ET), we examined the effect of ET on activities of extracellular signal-regulated kinases (ERKs) in rat aortic smooth muscle cells (SMCs). Treatment of rat aortic SMCs with ET-1 increased kinase activities toward myelin basic protein (MBP). Both 43- and 41-kDa proteins were activated when kinase assays were done in MBP-containing polyacrylamide gels after SDS-PAGE. These proteins were identified as ERK1 and ERK2 with immunoprecipitation and immunoblotting using antipeptide antibodies, respectively. These results indicate that ERKs mediate signal transduction by ET.

The effects of the selective ROCK inhibitor, Y27632, on ET-1-induced hypertrophic response in neonatal rat cardiac myocytes--possible involvement of Rho/ROCK pathway in cardiac muscle cell hypertrophy.

A small GTPase, Rho, participates in agonist-induced cytoskeletal organization and gene expression in many cell types including cardiac myocytes. However, little is known about the functions of Rho's downstream targets in cardiac myocytes. We examined the role of ROCK, a downstream target of Rho, in ET-1-induced hypertrophic response. Y27632, a selective ROCK inhibitor, inhibited ET-1-induced increases in natriuretic peptide production, cell size, protein synthesis, and myofibrillar organization. In addition, a dominant-negative mutant of p160ROCK suppressed ET-1-induced transcription of the BNP gene. These findings suggest that the Rho/ROCK pathway is an important component of ET-1-induced hypertrophic signals in cardiac myocytes.

Regulation of secretion and clearance of C-type natriuretic peptide in the interaction of vascular endothelial cells and smooth muscle cells.

OBJECTIVE: To clarify the significance of C-type natriuretic peptide in the interaction between endothelial cells and vascular smooth muscle cells by investigating the endothelial production of C-type natriuretic peptide and the clearance mechanism of C-type natriuretic peptide using the endothelial cells-smooth muscle cells co-culture system. RESULTS: Secretion of C-type natriuretic peptide in the direct co-culture of endothelial cells with smooth muscle cells elicited as much as a 60-fold increase compared with endothelial cells alone. The accumulation of intracellular cyclic GMP in the co-culture was consequently increased and the elevation of cyclic GMP level in the co-culture was abolished by the anti-C-type natriuretic peptide monoclonal antibody. The elevated cyclic GMP production in the co-culture was abolished by the anti-transforming growth factor-beta neutralizing antibody. Candoxatrilat (10(-6)-10(-4) mol/l), a neutral endopeptidase inhibitor, dose-dependently increased the concentrations of C-type natriuretic peptide in the culture medium with endothelial cells alone, but not in the endothelial cells-smooth muscle cells co-culture. The transcript of neutral endopeptidase messenger RNA was detected in endothelial cells but not in smooth muscle cells by reverse transcriptase polymerase chain reaction. Treatment with C-atrial natriuretic factor4-23 (10(-9)-10(-6) mol/l), the specific ligand for the clearance receptor of the natriuretic peptides, resulted in dose-dependent augmentation of C-type natriuretic peptide concentration and concomitant intracellular cyclic GMP production in the endothelial cells-smooth muscle cells co-culture but not in endothelial cells alone. CONCLUSION: The present study demonstrated that direct interaction between endothelial cells and smooth muscle cells augments C-type natriuretic peptide secretion from endothelial cells through transforming growth factor-beta activation, and revealed that the enzymatic degradation is responsible for the steady state level of C-type natriuretic peptide in endothelial cells alone and that the receptor-mediated clearance mainly determines the augmented level of C-type natriuretic peptide in the interaction between endothelial cells and smooth muscle cells. The results taken together raise the possibility that endothelial C-type natriuretic peptide might play a role in regulation of vascular tone and remodelling.

Lack of association between T-786-->C mutation in the 5'-flanking region of the endothelial nitric oxide synthase gene and essential hypertension.

Accumulating evidence strongly suggests that an alteration in nitric oxide metabolism is involved in the pathogenesis of hypertension. We recently found 2 polymorphisms in the endothelial nitric oxide synthase (eNOS) gene, a Glu298Asp missense variant in exon 7 and a T-786-->C variant in the 5'-flanking region, which are not linked to each other. In our previous reports, we showed a positive association between the Glu298Asp variant and essential hypertension, myocardial infarction, and coronary spastic angina. We also revealed that the T-786-->C variant is strongly associated with coronary spastic angina and leads to the reduction of the eNOS gene promoter activity. To further investigate the genetic involvement of the eNOS gene in essential hypertension, we examined the frequency of T-786-->C variant in two independent populations of persons with essential hypertension in Kyoto (n=215) and Kumamoto (n=186) and compared the frequency with that in each age- and gender-matched control (233 controls in Kyoto and 223 controls in Kumamoto). In both groups, the frequency of T-786-->C variant was similar in patients with hypertension and normal controls. In conclusion, the T-786-->C variant was not positively associated with essential hypertension. Given the evidence of positive association of another polymorphism in the eNOS gene, the Glu298Asp polymorphism, with essential hypertension, special attention will be required to interpret the results of a case-control study for genetic risk factors.

Lymphomatous polyarthritis in patients with peripheral T-cell lymphoma.

Direct involvement of the joints is a rare complication of malignant lymphoma and lymphoma cells in synovium or synovial fluid have been characterized in only a very few cases. We report two cases of CD4-positive, HTLV-I-negative peripheral T-cell lymphomas that manifested polyarthritis infiltrated with lymphoma cells which we further characterized. Patient 1, with a prior 7-year history of cutaneous T-cell lymphoma (mycosis fungoides) and polyarthralgia, developed pain and swelling in the right knee joint and right femoral region. Patient 2 was initially diagnosed with immunoblastic lymphadenopathy, later rediagnosed as the prodromal stage of T zone lymphoma. Seven years later she developed skin eruptions, cervical lymph node swelling, polyarthritis, and pleural effusion. Synovial fluid analysis in both cases showed predominant CD3+ or cytoplasmic CD3+, CD4+, and CD8- atypical lymphoid cell infiltration. In both cases the T-cell receptor beta and gamma chains were rearranged in the synovial fluid mononuclear cells. Analysis of these two cases and a review of the literature suggest that lymphoma cell infiltration of synovium occurs preferentially in patients with CD4+ peripheral T-cell lymphoma.

Cloacal inoculation with the Connecticut strain of avian infectious bronchitis virus: an attempt to produce nephropathogenic virus by in vivo passage using cloacal inoculation.

Avian infectious bronchitis virus (IBV) strain Connecticut A-5968 isolated from respiratory tissue of chickens in USA in the 1960s, is considered as representative of respiratory disease causing IBV strains. Specific pathogen free chicks inoculated with the strain via the cloaca replicated the virus more rapidly in their kidneys than did chicks inoculated with the same virus intratracheally. Virus passaged thirteen-times via the cloaca caused stronger nephrotropism and nephropathogenicity than the parent virus. It is suggested that cloacal inoculation of IBV provides new and interesting information concerning the mechanisms of nephropathogenicity of IBV which has became increasingly important worldwide during the last ten years.

Vasodilator therapy with hydralazine induces angiotensin AT receptor-mediated cardiomyocyte growth in mice lacking guanylyl cyclase-A.

BACKGROUND AND PURPOSE: Recent clinical guidelines advocate the use of the isosorbide dinitrate/hydralazine combination in treatment for heart failure. However, clinical and laboratory evidence suggest that some vasodilators may induce cardiac hypertrophy under uncertain conditions. This study investigated the effects and underlying mechanism of action of the vasodilator hydralazine on cardiac growth. EXPERIMENTAL APPROACH: Wild-type mice and animals deficient in guanylyl cyclase-A (GCA) and/or angiotensin receptors (AT(1) and AT(2) subtypes) were treated with hydralazine ( approximately 24 mg.kg(-1).day(-1) in drinking water) for 5 weeks. Cardiac mass and/or cardiomyocyte cross-sectional area, fibrosis (van Giessen-staining) and cardiac gene expression (real-time RT-PCR) were measured. KEY RESULTS: Hydralazine lowered blood pressure in mice of all genotypes. However, this treatment increased the heart and left ventricular to body weight ratios, as well as cardiomyocyte cross-sectional area, and cardiac expression of atrial natriuretic peptide mRNA in mice lacking GCA. Hydralazine did not affect cardiac hypertrophy in wild-type mice and mice lacking either AT(1) or AT(2) receptors alone. However, the pro-hypertrophic effect of hydralazine was prevented in mice lacking both GCA and AT(2), but not GCA and AT(1) receptors. However, hydralazine did decrease cardiac collagen deposition and collagen I mRNA (signs of cardiac fibrosis) in mice that were deficient in GCA, or both GCA and AT(2) receptors. CONCLUSIONS AND IMPLICATIONS: The vasodilator hydralazine induced AT(2) receptor-mediated cardiomyocyte growth under conditions of GCA deficiency. However, attenuation of cardiac fibrosis by hydralazine could be beneficial in the management of cardiac diseases.