Renal blood flow measurement with contrast-enhanced harmonic ultrasonography: evaluation of dopamine-induced changes in renal cortical perfusion in humans.

BACKGROUND: An accessible non-invasive method for evaluating renal regional blood flow in real time is highly desirable in the clinical setting. Recent progress in ultrasonography with microbubble contrast has allowed quantification of regional blood flow in animal models. AIMS: Goal ofthis study was to establish a convenient contrast--enhanced harmonic ultrasonography (CEHU) method for evaluating renal cortical blood flow in humans. METHODS: We carried out intermittent second harmonic imaging in 9 healthy volunteers. Pulse interval was progressively decreased from 4 s - 0.2 s during continuous venous infusion of the microbubble contrast agent. RESULTS: Pulse interval versus CEHU-derived acoustic intensity plots provided microbubble velocity (MV) and fractional vascular volume (FVV) during renal cortical perfusion in humans. Low-dose dopamine infusion (2 microg/min/kg) resulted in a significant increase in MV which correlated well with the increase in total renal blood flow (RBF) determined by a conventional study of p-aminohippurate clearance (C(PAH)) (r = 0.956, p < 0.0001). Although FVV was not significantly increased, alterations in CEHU-derived renal cortical blood flow calculated by the products of MV and FVV were also correlated with alterations in total RBF (r = 0.969, p < 0.0001). Thus, low-dose dopamine infusion increases renal cortical blood flow observed in CEHU, mainly by increasing MV. CONCLUSIONS: The present study shows that renal cortical blood flow in humans can be measured non-invasively by CEHU and that CEHU can be used for quantitatively evaluating changes induced by a therapeutic agent such as dopamine in flow velocity and in FVV.

Angiotensin II signaling and HB-EGF shedding via metalloproteinase in glomerular mesangial cells.

BACKGROUND: Angiotensin II (Ang II) has been implicated in the development of glomerulosclerosis by stimulating fibronectin (FN) synthesis. The processing and release of heparin binding-endothelin growth factor (HB-EGF) are activated by protein kinase C (PKC) and Ca2+ signaling. We studied the roles of HB-EGF and endothelial growth factor (EGF) receptor (EGFR) in Ang II-induced FN expression using mesangial cells. METHODS: Mesangial cells were prepared from mouse kidneys by the explant method and cells were used at passages 4 and 5. RESULTS: Ang II stimulated FN mRNA levels dose-dependently with a maximal increase (3.4-fold) after 12 hours of incubation. This action was completely inhibited by PKC inhibitors and slightly blocked by Ca2+ chelating agents. FN mRNA accumulation by Ang II was abolished by tyrosine kinase inhibitors, a specific inhibitor for EGFR (AG1478) and extracellular signal-regulated kinase (ERK) inactivation. Addition of neutralizing anti-HB-EGF antibody, as well as pretreatment with heparin or the metalloproteinase inhibitor batimastat abolished induction of FN expression by Ang II. In mesangial cells stably transfected with a chimeric construct containing HB-EGF and alkaline phosphatase (ALP) genes, ALP activity in incubation medium was rapidly increased by Ang II (1.7-fold at 0.5 min) and reached a 4.1-fold increase at two minutes. Ang II phosphorylated EGFR (maximal at 2 min) and ERK (maximal at 8 min) in a PKC- and metalloproteinase-dependent manner. Ang II stimulated the expression and release of transforming growth factor-beta (TGF-beta) via EGFR-mediated signaling, and the released TGF-beta also contributed to Ang II-mediated FN expression via EGFR transactivation. CONCLUSIONS: Ang II-mediated FN expression was regulated by autocrine effects of HB-EGF and TGF-beta, suggesting a novel paradigm for cross-talk between Ang II and growth factor receptor signaling pathways.

Angiotensin II type 2 receptor inhibits epidermal growth factor receptor transactivation by increasing association of SHP-1 tyrosine phosphatase.

Angiotensin (Ang) II has 2 major receptor isoforms, Ang type 1 (AT(1)) and Ang type (AT(2)). AT(1) transphosphorylates epidermal growth factor receptor (EGFR) to activate extracellular signal-regulated kinase (ERK). Although AT(2) was shown to inactivate ERK, the action of AT(2) on EGFR activation remains undefined. Using AT(2)-overexpressing vascular smooth muscle cells from AT(2) transgenic mice, we studied these undefined actions of AT(2). Maximal ERK activity induced by Ang II was increased 1.9- and 2.2-fold by AT(2) inhibition, which was abolished by orthovanadate but not okadaic acid or pertussis toxin. AT(2) inhibited AT(1)-mediated EGFR tyrosine phosphorylation by 63%. The activity of SHP-1 tyrosine phosphatase was significantly upregulated 1 minute after AT(2) stimulation and association of SHP-1 with EGFR was increased, whereas AT(2) failed to tyrosine phosphorylate SHP-1. Stable overexpression of SHP-1-dominant negative mutant completely abolished AT(2)-mediated inhibition of EGFR and ERK activation. AT(1)-mediated c-fos mRNA accumulation was attenuated by 48% by AT(2) stimulation. Induction of fibronectin gene containing an AP-1 responsive element in its 5'-flanking region was decreased by 37% after AT(2) stimulation, corresponding to the results of gel mobility assay with the AP-1 sequence of fibronectin as a probe. These findings suggested that AT(2) inhibits ERK activity by inducing SHP-1 activity, leading to decreases in AP-1 activity and AP-1-regulated gene expression, in which EGFR dephosphorylation plays an important role via association of SHP-1.

Safety and availability of doxazosin in treating hypertensive patients with chronic renal failure.

Hypertension accelerates the progression of renal disease in patients with chronic renal failure. Doxazosin, an alpha1-antagonist, is an antihypertensive agent with a long half-life. In this study, 15 patients with chronic renal failure were treated only with doxazosin and diuretics for 6 months and their blood pressure, renal parameters and lipid profile were measured. The initial dose of doxazosin was 2 mg/day and it was titrated until blood pressure was normalized. The average dose was 5.6 mg/day. As expected, systolic and diastolic blood pressure were decreased with treatment (165/91 mmHg to 135/73 mmHg). The drop in blood pressure was associated with an increase in glomerular filtration and a decrease in plasma BUN and creatinine levels. Reduction in mean blood pressure and decrease in proteinuria had a significant positive correlation (r=0.048, p=0.007). Proteinuria was decreased from 1.8 mg/day to 1.3 mg/day with doxazosin treatment and triglycerides also decreased, while HDL-cholesterol was increased. No side effects were observed. These results indicate that doxazosin is an efficient depressor agent with renal protective actions and that higher doses of doxazosin can be safely given to patients with chronic renal failure.

Implantation of bone marrow mononuclear cells into ischemic myocardium enhances collateral perfusion and regional function via side supply of angioblasts, angiogenic ligands, and cytokines.

BACKGROUND: Bone marrow implantation (BMI) was shown to enhance angiogenesis in a rat ischemic heart model. This preclinical study using a swine model was designed to test the safety and therapeutic effectiveness of BMI. METHODS AND RESULTS: BM-derived mononuclear cells (BM-MNCs) were injected into a zone made ischemic by coronary artery ligation. Three weeks after BMI, regional blood flow and capillary densities were significantly higher (4.6- and 2.8-fold, respectively), and cardiac function was improved. Angiography revealed that there was a marked increase (5.7-fold) in number of visible collateral vessels. Implantation of porcine coronary microvascular endothelial cells (CMECs) did not cause any significant increase in capillary densities. Labeled BM-MNCs were incorporated into approximately 31% of neocapillaries and corresponded to approximately 8.7% of macrophages but did not actively survive as myoblasts or fibroblasts. There was no bone formation by osteoblasts or malignant ventricular arrhythmia. Time-dependent changes in plasma levels for cardiac enzymes (troponin I and creatine kinase-MB) did not differ between the BMI, CMEC, and medium-alone implantation groups. BM-MNCs contained 16% of endothelial-lineage cells and expressed basic fibroblast growth factor>>vascular endothelial growth factor>angiopoietin 1 mRNAs, and their cardiac levels were significantly upregulated by BMI. Cardiac interleukin-1beta and tumor necrosis factor-alpha mRNA expression were also induced by BMI but not by CMEC implantation. BM-MNCs were actively differentiated to endothelial cells in vitro and formed network structure with human umbilical vein endothelial cells. CONCLUSIONS: BMI may constitute a novel safety strategy for achieving optimal therapeutic angiogenesis by the natural ability of the BM cells to secrete potent angiogenic ligands and cytokines as well as to be incorporated into foci of neovascularization.

Effect of angiotensin II type 2 receptor on tyrosine kinase Pyk2 and c-Jun NH2-terminal kinase via SHP-1 tyrosine phosphatase activity: evidence from vascular-targeted transgenic mice of AT2 receptor.

Angiotensin II (Ang II) has two major receptor isoforms, AT1 and AT2. AT1 transphosphorylates Ca(2+)-sensitive tyrosine kinase Pyk2 to activate c-Jun NH2-terminal kinase (JNK). Although AT2 inactivates extracellular signal-regulated kinase (ERK) via tyrosine phosphatases (PTP), the action of AT2 on Pyk2 and JNK remains undefined. Using AT2-overexpressing vascular smooth muscle cells (AT2-VSMC) from AT2-transgenic mice, we studied these undefined actions of AT2. AT1-mediated JNK activity was increased 2.2-fold by AT2 inhibition, which was abolished by orthovanadate. AT2 did not affect AT1-mediated Pyk2 phosphorylation, but attenuated c-Jun mRNA accumulation by 32%. The activity of src-homology 2 domain-containing PTP (SHP-1) was significantly upregulated 1 min after AT2 stimulation. Stable overexpression of SHP-1 dominant negative mutant in AT2-VSMC completely abolished AT2-mediated inhibition of JNK activation and c-Jun expression. These findings suggest that AT2 inhibits JNK activity by affecting the downstream signal of Pyk2 in a SHP-1-dependent manner, leading to a decrease in c-Jun expression.

Angiotensin AT(1) and AT(2) receptors differentially regulate angiopoietin-2 and vascular endothelial growth factor expression and angiogenesis by modulating heparin binding-epidermal growth factor (EGF)-mediated EGF receptor transactivation.

Angiotensin II (Ang II)-mediated signals are transmitted via heparin binding epidermal growth factor (EGF)-like growth factor (HB-EGF) release followed by transactivation of EGF receptor (EGFR). Although Ang II and HB-EGF induce angiogenesis, their link to the angiopoietin (Ang)-Tie2 system remains undefined. We tested the effects of Ang II on Ang1, Ang2, or Tie2 expression in cardiac microvascular endothelial cells expressing the Ang II receptors AT(1) and AT(2). Ang II significantly induced Ang2 mRNA accumulations without affecting Ang1 or Tie2 expression, which was inhibited by protein kinase C inhibitors and by intracellular Ca(2+) chelating agents. Ang II transactivated EGFR via AT(1), and inhibition of EGFR abolished the induction of Ang2. Ang II caused processing of pro-HB-EGF in a metalloproteinase-dependent manner to stimulate maturation and release of HB-EGF. Neutralizing anti-HB-EGF antibody blocked EGFR phosphorylation by Ang II. Ang II also upregulated vascular endothelial growth factor (VEGF) expression in an HB-EGF/EGFR-dependent manner. AT(2) inhibited AT(1)-mediated Ang2 expression and phosphorylation of EGFR. In an in vivo corneal assay, AT(1) induced angiogenesis in an HB-EGF-dependent manner and enhanced the angiogenic activity of VEGF. Although neither Ang2 nor Ang1 alone induced angiogenesis, soluble Tie2-Fc that binds to angiopoietins attenuated AT(1)-mediated angiogenesis. These findings suggested that (1) Ang II induces Ang2 and VEGF expression without affecting Ang1 or Tie2 and (2) AT(1) stimulates processing of pro-HB-EGF by metalloproteinases, and the released HB-EGF transactivates EGFR to induce angiogenesis via the combined effect of Ang2 and VEGF, whereas AT(2) attenuates them by blocking EGFR phosphorylation. Thus, Ang II is involved in the VEGF-Ang-Tie2 system via HB-EGF-mediated EGFR transactivation, and this link should be considerable in pathological conditions in which collateral blood flow is required.

Multiple spontaneous small bowel perforations due to systemic cholesterol atheromatous embolism.

A-65-year-old man was admitted for coronary and peripheral angiography to evaluate angina pectoris and peripheral vascular disease. Following angiography, he suffered from blue toes, livedo reticularis and progressive renal failure. The patient's condition continued to deteriorate, including the development of malnutrition. Four months later he suddenly developed panperitonitis, went into shock and died. The autopsy verified multiple perforations of the small bowel with disseminated cholesterol atheromatous embolism. The other organs including kidney were also invaded by atheroembolism. This was a rare case of multiple spontaneous perforations of small bowel due to systemic cholesterol atheromatous embolism.

[Aortic valvular vegetation in annuloaortic ectasia due to idiopathic cystic medial necrosis: a case report].

Annuloaortic ectasia is often accompanied by Marfan syndrome and associated with infective endocarditis usually involving the mitral valves. We treated a patient with annuloaortic ectasia due to idiopathic cystic medial necrosis who developed congestive heart failure with aortic valvular vegetation. A 56-year-old man had dyspnea on effort since the beginning of January, 1997 and was admitted to our hospital on April 6, 1997 because of orthopnea. The diagnosis was congestive heart failure due to severe aortic regurgitation with annuloaortic ectasia detected by echocardiography. Medication and rest after hospitalization relieved his symptoms but congestive heart failure deteriorated after he had a high fever. At this time, a vegetation attached to the noncoronary cusp of the aortic valve was found which had not been detected on admission. Blood culture yielded Streptococcus sanguis. The diagnosis was infective endocarditis involving the aortic valve. Surgical correction (Bentall method) improved congestive heart failure and he was discharged on August 4, 1997 without recurrence of endocarditis. Infective endocarditis involving the aortic valves is a possible cause of development or deterioration of congestive heart failure in patients with annuloaortic ectasia.