Abrogation of protein phosphatase 6 promotes skin carcinogenesis induced by DMBA.

Somatic mutations in the gene encoding the catalytic subunit of protein phosphatase 6 (Ppp6c) have been identified in malignant melanoma and are thought to function as a driver in B-raf- or N-ras-driven tumorigenesis. To assess the role of Ppp6c in carcinogenesis, we generated skin keratinocyte-specific Ppp6c conditional knockout mice and performed two-stage skin carcinogenesis analysis. Ppp6c deficiency induced papilloma formation with 7,12-dimethylbenz (a) anthracene (DMBA) only, and development of those papillomas was significantly accelerated compared with that seen following DMBA/TPA (12-O-tetradecanoylphorbol 13-acetate) treatment of wild-type mice. NF-κB activation either by tumor necrosis factor (TNF)-α or interleukin (IL)-1ß was enhanced in Ppp6c-deficient keratinocytes. Overall, we conclude that Ppp6c deficiency predisposes mice to skin carcinogenesis initiated by DMBA. This is the first report showing that such deficiency promotes tumor formation in mice.

Rat neuronal leucine-rich repeat protein-3: cloning and regulation of the gene expression.

Rat neuronal leucine-rich repeat protein-3 (rNLRR-3) gene was isolated and cloned from fibrosarcoma cells overexpressing c-Ha-ras. Stable expression of constitutively active forms of Ras (H-Ras(V12) or v-H-Ras) led to a two- to fourfold increase in rNLRR-3 mRNA in rat normal fibroblasts (3Y1). When cells expressing H-Ras(V12) were treated with mitogen activated protein kinase (MAPK) kinase inhibitors (U0126, PD98059), suppression of rNLRR-3 mRNA correlated well with a reduction in MAPK activity. Epidermal growth factor (EGF) led to elevation of rNLRR-3 gene expression about 4 h after stimulation of normal fibroblasts. U0126 completely suppressed the induction by EGF of rNLRR-3 mRNA with abrogation of MAPK phosphorylation. U0126 inhibited the basal transcription of rNLRR-3. LY294002, a PI3 kinase inhibitor, showed a lesser effect on expression of the gene. These results indicate that rNLRR-3 gene expression is regulated mainly through the Ras-MAPK signaling pathway in fibroblasts.

Device-based change in left ventricular shape: a new concept for the treatment of dilated cardiomyopathy.

OBJECTIVE: We tested a unique new device, the Myosplint device (Myocor, Inc, Maple Grove, Minn), which is designed to change left ventricular shape, reduce left ventricular wall stress, and improve left ventricular systolic function. METHODS: Heart failure was induced in 15 dogs over 27 days by rapid pacing (230 beats/min). Seven animals underwent sham surgery, and 8 animals received 3 transventricular Myosplint devices each. Myosplint devices were tightened to create a symmetric bilobular left ventricular shape and were adjusted to produce a calculated 20% reduction in wall stress. Hemodynamic, 2-dimensional, and 3-dimensional echocardiographic studies were recorded at baseline, immediately after Myosplint placement (acute change), and at 1 month after both groups had a reduced rate (190 beats/min) of pacing designed to maintain heart failure. RESULTS: The Myosplint group had significant sustained improvements in left ventricular ejection fraction from baseline, to the acute change, to 1 month (19% +/- 5%; 36% +/- 8%; 39% +/- 13%) and reductions of left ventricular end-systolic volumes (73 +/- 9 mL; 34 +/- 5 mL; 42 +/- 12 mL) and end-systolic wall stress by 39% (341 +/- 68 10(3) dynes x cm(- 2) to 206 +/- 28 10(3) dynes x cm(-2)) acutely and 31% (372 +/- 83 10(3) dynes x cm(-2) to 250 +/- 40 10(3) dynes x cm(-2)) at 1 month. There were no significant changes in mitral regurgitation. CONCLUSION: Application of a Myosplint device to a dilated impaired left ventricle resulted in reduced wall stress and improved left ventricular systolic function that was sustained at 1 month. Device-based shape change is a promising new opportunity to treat patients with dilated cardiomyopathy.

In vivo hemodynamic performance of the Cleveland Clinic CorAide blood pump in calves.

BACKGROUND: The Cleveland Clinic CorAide left ventricular assist system is based on a small implantable continuous-flow centrifugal blood pump with a completely suspended rotating assembly designed for long-term circulatory support (5 to 10 years). METHODS: Between June 1999 and August 2000, the CorAide blood pump was implanted in 10 calves for 1 month and in 3 calves for 3 months. RESULTS: The mean pump flow and arterial pressure were 6.1 +/- 1.1 L/min and 97 +/- 5 mm Hg, respectively. The mean plasma free-hemoglobin level after postoperative day 3 was 2.0 +/- 1.8 mg/dL. Renal and hepatic function remained normal in all cases. There was no incidence of mechanical failure, hemolysis, bleeding, or systemic organ dysfunction in any of the cases. Significant findings at autopsy were limited to two cases of renal infarction, one of which was associated with an outflow graft infection. CONCLUSIONS: The CorAide blood pump is easily implanted, reliable, nonhemolytic, and nonthrombogenic, positioning it as a leading third-generation, continuous-flow left ventricular assist system with a completely suspended rotor.

[Two cases of methicillin-resistant Staphylococcus aureus (MRSA) sepsis following craniotomy].

We report here two cases of MRSA sepsis following craniotomy. In case 1, a petroclival meningioma was subtotally removed and lumbar drainage was inserted postoperatively to prevent cerebrospinal fluid leakage. Ventriculo-peritoneal shunt was performed after meningitis was treated with vancomycin and panipenem/betamipron. Two weeks after the procedure, the patient revealed continuous spiking fevers related to MRSA sepsis, which did not improve with vancomycin and arbekacin administration. The focus of infection was found by scintigraphy and CT by 67Ga to be spondylo-diskitis at the level of L2-L3. The lesion was removed and bone from the iliac crest grafted. In case 2, seven days after surgery for multiple meningioma, the patient exhibited spiking fevers and swelling in the left leg. The central venous catheter was removed from the left femoral vein and MRSA was found from blood culture. The patient was treated with arbekacin (200 mg/day). Venous thrombosis diagnosed by CT was treated with heparin. Symptoms related to the infection and laboratory data did not improve because the concentration of arbekacin in the blood did not reach an effective level. The symptoms markedly improved when the dose of arbekacin was doubled (400 mg/day).

A possible mechanism of TPA-mediated downregulation of neurotrophin-3 gene expression in rat cultured vascular smooth muscle cells.

We have previously reported that in cultured rat vascular smooth muscle cells (VSMCs), neurotrophin-3 (NT-3) gene expression was suppressed by TPA (12-O-tetradecanoyl phorbol-13-acetate), which induces an AP-1 transcription factor. In the present study, to clarify the mechanism for TPA-mediated downregulation of NT-3 gene expression, effects of cycloheximide and dexamethasone (Dex) on the TPA-mediated downregulation were examined in VSMCs. Pretreatment with cycloheximide, an inhibitor of protein synthesis, or simultaneous treatment with Dex, an inhibitor of AP-1, suppressed the TPA-mediated downregulation of NT-3 gene expression. Furthermore, co-transfection of c-fos and c-jun expression vectors into VSMCs resulted in decrease in the NT-3 gene expression. The present findings suggest that TPA-induced AP-1 de novo synthesis causes the downregulation of NT-3 gene expression in VSMCs.

Preoperative risk factors for right ventricular failure after implantable left ventricular assist device insertion.

BACKGROUND: Implantable left ventricular assist device (LVAD) insertion complicated by early right ventricular (RV) failure has a poor prognosis and is generally unpredictable. METHODS: To determine preoperative risk factors for perioperative RV failure after LVAD insertion, patient characteristics and preoperative hemodynamics were analyzed in 100 patients with the HeartMate LVAD (Thermo Cardiosystems, Inc, Woburn, MA) at the Cleveland Clinic. RESULTS: RV assist device support was required for 11 patients (RVAD group). RVAD use was significantly higher in younger patients, female patients, smaller patients, and myocarditis patients. There was no significant difference in the cardiac index, RV ejection fraction, or right atrial pressure between the two groups preoperatively. The preoperative mean pulmonary arterial pressure (PAP) and RV stroke work index (RV SWI) were significantly lower in the RVAD group (p = 0.015 and p = 0.011, respectively). Survival to transplant was poor in the RVAD group (27%) and was 83% in the no-RVAD group. CONCLUSIONS: The need for perioperative RVAD support was low, only 11%. Preoperative low PAP and low RV SWI were significant risk factors for RVAD use.

The Cleveland Clinic-Nimbus total artificial heart. In vivo hemodynamic performance in calves and preclinical studies.

In vitro function of the Cleveland Clinic-Nimbus electrohydraulic total artificial heart met National Heart, Lung, and Blood Institute hemodynamic guidelines for such devices. In a series of in vivo experiments, we implanted the total artificial heart in eight calves (mean weight 87 kg), one for a short-term experiment and seven for long-term experiments. The mean blood flow during support was 7.7 +/- 1.6 L/min with left atrial pressure 13 +/- 6 mm Hg, right atrial pressure 13 +/- 4 mm Hg, and aortic pressure 97 +/- 9 mm hg. Maximum pump flow (9.6 L/min) occurred after 4 days of support as a result of the high resting cardiac output of the animals. A 10% to 15% right pump stroke-volume limit effectively balanced atrial pressures, and afterload insensitivity was confirmed by the in vivo studies. Calves tolerated treadmill exercise studies well, with an average duration of 22 minutes and an average top speed of 2.1 mph. The experiments were terminated after 1 day to 120 days of support (mean 32 days). Most experiments were terminated as a result of correctable mechanical problems. In a separate study of six adult human patients undergoing orthotopic cardiac transplantation, five showed an excellent fit for the Cleveland Clinic-Nimbus total artificial heart. Further studies using chest roentgenograms, chest measurements, and transesophageal echocardiography should help predict fit of the total artificial heart in potential candidates. Initial candidates for a "vented-electric" version of the Cleveland Clinic-Nimbus total artificial heart are patients for whom univentricular (left ventricular assist device) support is not appropriate, but who require mechanical support as a bridge to cardiac transplantation.

The effects of radiation therapy on the tissue capsule of soft tissue implants.

A prosthesis has been developed for cosmesis after lumpectomy surgery for breast carcinoma. The device is saline filled and percutaneously adjustable in volume to permit an optimal cosmetic result after surgical wound healing. A series of 24 studies of 18 weeks' duration using the adult rabbit animal model were first used to study tissue capsule formation around textured versus smooth surface control implants and to evaluate the effects of volume adjustments on the tissue capsule. Single or multiple adjustments of implant volume had no effect on tissue capsule thickness or morphology. Because lumpectomy surgery is invariably followed by radiation therapy, a series of six studies was then conducted to determine the effects of a typical course of radiation therapy on tissue capsule formation. One week after device implantation, a 4 x 4 cm field including the implant was irradiated with 5,000 rad (200 rad/day x 5 days/week x 5 weeks). The animals were maintained for a 6 week period after radiation treatment. After sacrifice, the implants were removed, and the tissue capsules studied using conventional histologic techniques, including scanning and transmission electron microscopy. There was no statistically significant difference in tissue capsule thickness compared to nonirradiated controls. Tissue capsule morphology, however, differed markedly. Radiation therapy decreased angiogenesis, cellularity, and the inflammatory cell response to the implants. Qualitatively, radiation treatment seemingly improved rather than compromised the connective tissue response to the implants.

Effects of preserving mitral apparatus on ventricular systolic function in mitral valve operations in dogs.

The mitral apparatus can affect left ventricular function through various mechanisms, such as the direct mechanical coupling between the mitral anulus and papillary muscle and the mitral annular contraction. To evaluate the relative contribution of these mechanisms, we investigated in five groups of 35 dogs that underwent open chest operations how preservation of the mitral apparatus affects left ventricular systolic function. We performed atriotomy in the first group. We sutured a prosthetic rigid ring around the mitral anulus in the second group. We replaced the mitral valve and preserved the anterior chordae in the third group, the posterior chordae in the fourth group, and no chordae in the fifth group. The postoperative percentage of recovery of ventricular function (as assessed by the slope of the end-systolic pressure-volume relation) from preoperative control values were 92.2% +/- 4.8%, 89.5% +/- 12.8%, 85.7% +/- 9.5%, 75.1% +/- 12.9%, and 61.3% +/- 8.0%, respectively. Preservation of the mitral apparatus significantly improved left ventricular function compared with that of conventional mitral valve replacement. The average relative contribution of the ventricular muscle to left ventricular function, the mitral anulus-papillary muscle continuity, and the mitral annular contraction were 66.5%, 30.6%, and 2.9%, respectively. We conclude that it is more beneficial to preserve the mitral apparatus in mitral valve replacement. The prosthetic ring does not detract from the functional benefit of the preservation of the mitral apparatus.

Development of a magnetically operated artificial urethral sphincter. Chronic effects of compression on the skin structure and blood flow.

The artificial urethral sphincter (AUS) has been in clinical use for more than 20 years. Currently available AUS devices, however, are difficult to use and not entirely reliable. A magnetically operated AUS is currently under the development. Although the skin between the magnets will be compressed all day long, little information exists on the effects of chronic pressure on the skin structure and blood flow. In five miniature pigs, two internal magnets and one control metal disk were implanted subcutaneously at three different positions, and external magnets with differing magnetic forces were applied to the skin overlying the internal magnets for six weeks. In four pigs, the skin blood flow was measured by a laser Doppler flow meter applying different pressures. Compression of 10 mmHg preserved normal skin morphology in all but one animal where blood flow had not recovered 2 weeks after surgery. Compression of 20 mmHg for 6 weeks, however, produced pressure ulcers in all five cases (p < 0.05 vs. 10 mmHg group). The skin blood flow declined for pressures exceeding 20 mmHg (0 mmHg: 4.3 +/- 1.2, 10 mmHg: 4.3 +/- 3.3, 20 mmHg: 2.6 +/- 2.7 ml/min/100 g). We concluded that the magnetically operated AUS should use a magnetic coupling with a pressure less than 10 mmHg exerted on the interposing skin.

Effects of diltiazem and noradrenaline on extracellular potassium changes in the globally ischaemic rat heart.

OBJECTIVE: The aim was to investigate the effects of a calcium antagonist (diltiazem) and a catecholamine (noradrenaline) on extracellular potassium accumulation during global ischaemia. METHODS: Extracellular potassium concentration ([K+]e) was measured during 30 min global ischaemia in the isolated rat heart using a valinomycin potassium sensitive electrode. Contracture development during ischaemia was measured throughout with an intraventricular balloon inserted into the left ventricle and myocardial adenine nucleotides were measured in separate series of hearts. RESULTS: In control hearts, [K+]e showed a characteristic triphasic change during 30 min global ischaemia. This consisted of an early rising phase followed by a transient falling phase after the initial peak of [K+]e, and then a late rising phase. Diltiazem suppressed the rate of rise of [K+]e during early ischaemia, but extended the time course of the early [K+]e rise with the higher dose, abolishing the transient falling phase of [K+]e. During late ischaemia, the rise in [K+]e was attenuated by diltiazem. Noradrenaline also suppressed the early extracellular potassium accumulation, but in contrast to diltiazem, hastened the time course of the late [K+]e rise. CONCLUSIONS: Although diltiazem suppresses the early potassium loss during ischaemia as previously described, the drug also decreases the [K+]e fall by some as yet unknown mechanism, so that the [K+]e level becomes higher than control during the falling phase.

Effects of mechanical ventilation and spontaneous respiration on hemodynamics in calves with total artificial hearts.

The effects of respiration on hemodynamics were evaluated in four Holstein calves with total artificial hearts (TAH). The electrohydraulic actuated E4T-TAH has a continuously reciprocating actuator packaged between two alternately ejecting blood pumps that passively fill. The hemodynamic parameters (right atrial pressure [RAP], left atrial pressure [LAP], pulmonary artery pressure [PAP], aortic pressure [AoP]), and right and left pump filling (Rt% fill and Lt% fill) were measured when the animal was intubated and mechanically ventilated. These measurements were repeated with spontaneous respiration after the animal was extubated. With mechanical ventilation, LAP, PAP, and AoP were significantly higher during inspiration than during expiration. However, RAP during inspiration was slightly lower than that during expiration. The Rt% fill during inspiration was significantly lower than during expiration, but Lt% fill during inspiration was significantly higher than during expiration. During spontaneous respiration, these changes were opposite to those observed during mechanical ventilation. That mechanical ventilation generates positive intrathoracic pressure during inspiration, but spontaneous respiration generates negative pressure may explain these results. The change in venous return to the right atrium caused the change in RAP to be opposite in direction to that of the other pressures.

Effects of glibenclamide and nicorandil on cardiac function during ischemia and reperfusion in isolated perfused rat hearts.

We examined influences of a blocker (glibenclamide) and an opener (nicorandil) of the ATP-sensitive potassium (KATP) channel on extracellular K concentration [( K+]e), as well as the myocardial function and metabolites during global ischemia and reperfusion in Langendorff-perfused rat heart preparation. In control hearts, [K+]e began to rise 20 s after the onset of ischemia up to an initial peak (8.3 +/- 0.3 mM) at 2.5 +/- 0.7 min, then fell to 6.0 +/- 0.8 mM after 8.2 +/- 0.7 min, and then rose progressively to 14.6 +/- 0.8 mM at the end of 30 min of ischemia. Glibenclamide (50 microM) reduced the initial peak of [K+]e to 7.2 +/- 0.3 mM (P less than 0.01), and nicorandil (200 microM) increased it to 9.4 +/- 0.6 mM (P less than 0.01). There were no significant differences in [K+]e values among all groups at the end of ischemia. During ischemia, nicorandil decreased the time to mechanical arrest from 1.9 +/- 0.1 min to 1.5 +/- 0.1 min, whereas it was increased by glibenclamide to 2.7 +/- 0.4 min. In control hearts, the time to onset of ischemic contracture was 14.7 +/- 1.8 min. Nicorandil delayed onset of contracture and glibenclamide accelerated it. Thus we have confirmed that some part of the early increase in [K+]e during ischemia is attributable to K+ efflux through the KATP channel in our model, and opening of the KATP channel may contribute to a rapid reduction of the contractility of the ischemic myocardium that subsequently protects the myocardium against further ischemic injury.

Beneficial effect of pericardial meshing on left ventricular pump performance in dogs.

To evaluate the effects of pericardial meshing (multiple incisions on the pericardium) on cardiac function, we examined left ventricular pump performance before and after pericardial meshing in six open chest dogs. We evaluated left ventricular systolic properties with the slope of end-systolic pressure-volume relation and diastolic properties with end-diastolic pressure-volume relation (chamber compliance). Overall left ventricular performance was assessed with end-diastolic pressure versus cardiac output relation. Left ventricular chamber compliance was increased (31.3%) with pericardial meshing compared with direct closure of the pericardium, and cardiac output was increased (26.7%) for any given left ventricular end-diastolic pressure. The slope of the end-systolic pressure-volume relation was not altered in pericardial meshing. These results suggest that pericardial meshing improves left ventricular pump performance as a result of increasing left ventricular chamber compliance. This technique may benefit cardiac pump performance that is depressed by the direct closure of the pericardium after cardiac operations.

Effects of left heart bypass on right ventricular performance. Evaluation of the right ventricular end-systolic and end-diastolic pressure-volume relation in the in situ normal canine heart.

Although left heart bypass has gained popularity as a powerful technique to assist the severely failed left heart, apparent right heart failure has often developed during the bypass procedure. We investigated whether the coexisting right heart failure is attributable to the left heart bypass in 16 open-chest dogs. We evaluated the effects of left heart bypass on the right ventricular systolic properties by the slope of the end-systolic pressure-volume relation and its effects on the diastolic properties by chamber compliance. Overall right ventricular performance was assessed by the end-diastolic pressure versus cardiac output relationship. The left heart bypass decreased the slope slightly when the assisted flow ratio exceeded 75% (-14% +/- 8% at the assisted flow ratio of 100%, p less than 0.02) and thus had a deleterious influence on right ventricular performance. The left heart bypass, on the other hand, had a counteracting beneficial influence on right ventricular performance through the increase in chamber compliance (38% +/- 5%, p less than 0.01) and the decrease in pulmonary arterial input resistance (-15% +/- 12%, p less than 0.01). The net effect of the left heart bypass was the increase in cardiac output (20% +/- 2%, p less than 0.05) for any given right ventricular end-diastolic pressure. We conclude that in normal hearts the left heart bypass augments right ventricular performance. We ascribe these beneficial effects to diastolic ventricular interdependence and afterload unloading.

A new method of retrograde cardioplegic administration. Right ventricular protection by right atrial perfusion cooling.

Retrograde administration of cardioplegic solution via the right atrium with continuous cooling of the right ventricular cavity (right atrial perfusion cooling) was assessed for its protective effect in 12 dogs with occlusion of the right coronary artery subjected to global ischemia for 60 minutes. After an initial administration of 4 degrees C crystalloid cardioplegic solution by antegrade aortic perfusion, myocardial protection was established either by right atrial perfusion cooling (group I; n = 6) or by antegrade aortic perfusion alone (group II; n = 6). The right ventricular temperature was approximately 15 degrees C in group I and 20 degrees C in group II. After ischemia for 60 minutes, the adenosine triphosphate content of the right ventricular free wall was significantly higher in group I than in group II (24.4 +/- 1.45 versus 13.8 +/- 2.34 mumol/gm dry weight, p less than 0.05). The percent recovery of right ventricular contractility, which was evaluated by end-systolic pressure-volume relationships, was significantly better in group I at each reperfusion period (30 minutes: 130.0% +/- 9.6% versus 86.1% +/- 11.8%, p less than 0.05; 60 minutes: 159.6% +/- 12.9% versus 96.5% +/- 20.1%, p less than 0.05). Postischemic right ventricular stiffness (reciprocal value of compliance) increased in group II compared with group I, although the difference was not statistically significant. There were no major differences in percent recovery of the left ventricular end-systolic pressure-volume relationships between the two groups. The evidence suggests that the right atrial perfusion cooling method produces excellent right ventricular protection.

The mechanism of protective effect of diltiazem on reperfusion-induced arrhythmias in isolated rat heart.

This investigation was undertaken to define the mechanism by which diltiazem protects against life-threatening, reperfusion-induced arrhythmias. Using an isolated retrogressively perfused rat heart preparation with transient coronary artery occlusion, we compared the effects of diltiazem in its active form (d-cis) to its stereo-isomer (1-cis). Pre-ischemic administration of d-diltiazem (5 x 10(-8), 5 x 10(-7), 5 x 10(-6) M) caused a dose-dependent reduction in ventricular arrhythmias upon reperfusion following 10 min of regional ischemia. The incidence of reperfusion-induced ventricular fibrillation (RVF) was 50%, 0% (p less than 0.05) and 0% (p less than 0.05) with 5 x 10(-8), 5 x 10(-7), 5 x 10(-6) M diltiazem, respectively, compared with 60% in the control group. Preischemic administration of the 1-isomer caused different dose-dependent reduction in RVF. With 5 x 10(-6) M, the 1-isomer also reduced the incidence of RVF to 0% (p less than 0.05). However below this concentration it was ineffective (67%). D-diltiazem (5 x 10(-7) and 5 x 10(-6) M) increased coronary flow from 11.5 +/- 1.9 ml/min to 15.3 +/- 1.6 ml/min (p less than 0.05) and 15.2 +/- 1.0 ml/min (p less than 0.05) respectively, prior to ischemia. In contrast, the same dose of the 1-isomer did not alter coronary flow. The highest dose (5 x 10(-6) M) of d-diltiazem decreased heart rate by approximately 30% during the reperfusion phase, but all other concentrations had no significant effects.(ABSTRACT TRUNCATED AT 250 WORDS)

[Clinical assessment of anticoagulation therapy during left ventricular assist with artificial heart: induction with protease inhibitor].

Left ventricular assist device was attached to five patients suffered from severe low cardiac output after open heart surgery. In two patients, anticoagulation therapy with heparin started just after the operation. Repeated operations for hemostasis were required because of massive bleeding in these two patients. Anticoagulation therapy was not performed in another one, and thrombus formation in the device was recognized in this patient. In the other two patients, anticoagulation therapy was started with large dose of protease inhibitor (gabexate mesilate or nafamstat mesilate). Heparin infusion was combined with protease inhibitor during the period of weaning from the device. Thrombosis and massive bleeding were not recognized in these two patients, and they were able to wean from the device successfully.