Adaptation of non-linear mixed amount with zero amount response surface model for analysis of concentration-dependent synergism and safety with midazolam, alfentanil, and propofol sedation.

BACKGROUND: The non-linear mixed amount with zero amounts response surface model can be used to describe drug interactions and predict loss of response to noxious stimuli and respiratory depression. We aimed to determine whether this response surface model could be used to model sedation with the triple drug combination of midazolam, alfentanil and propofol. METHODS: Sedation was monitored in 56 patients undergoing gastrointestinal endoscopy (modelling group) using modified alertness/sedation scores. A total of 227 combinations of effect-site concentrations were derived from pharmacokinetic models. Accuracy and the area under the receiver operating characteristic curve were calculated. Accuracy was defined as an absolute difference <0.5 between the binary patient responses and the predicted probability of loss of responsiveness. Validation was performed with a separate group (validation group) of 47 patients. RESULTS: Effect-site concentration ranged from 0 to 108 ng ml-1 for midazolam, 0-156 ng ml-1 for alfentanil, and 0-2.6 µg ml-1 for propofol in both groups. Synergy was strongest with midazolam and alfentanil (24.3% decrease in U50, concentration for half maximal drug effect). Adding propofol, a third drug, offered little additional synergy (25.8% decrease in U50). Two patients (3%) experienced respiratory depression. Model accuracy was 83% and 76%, area under the curve was 0.87 and 0.80 for the modelling and validation group, respectively. CONCLUSION: The non-linear mixed amount with zero amounts triple interaction response surface model predicts patient sedation responses during endoscopy with combinations of midazolam, alfentanil, or propofol that fall within clinical use. Our model also suggests a safety margin of alfentanil fraction <0.12 that avoids respiratory depression after loss of responsiveness.

An efficient transfection method for mouse embryonic stem cells.

Embryonic stem (ES) cells are considered to have potentials for tissue regeneration and treatment of diverse human diseases. ES cells are capable of indefinite renewal and proliferation, which can be induced to differentiate into tissues of all three germ lines. Despite these exciting potential, it remains unclear as to how the renewal and differentiation programs are operated and regulated at the genetic level. Genetic manipulation such as delivery of exogenous gene expression or knockdown with small interfering RNA (siRNA) is commonly used in most of cancer or transformed cells but relatively rare in ES cells. In this study, we compare the transfection efficacies of several liposome-based transfection methods by introduction of a plasmid encoding enhanced green fluorescent protein (EGFP) into mouse ES (mES) cells. Our results show that transfection by Effectene achieves the efficiency of >98% in CCE and >80% in D3 cells. The optimal ratio of DNA:Effectene for EGFP transfection is between 1:4 and 1:8. Transient-expressed EGFP or endogenous protein kinase A (PKA) were significantly knocked down by Effectene transfection of specific siRNA. High EGFP level expression and accumulation in mES cells induces minor cytotoxicity but can be reduced by introducing siRNA of EGFP. Further, this transfection method did not significantly affect mES properties of proliferation or differentiation. Our results provide an optimal protocol to achieve an efficient transfection for mES cells.

Inhibition of angiotensin II induced endothelin-1 gene expression by 17-beta-oestradiol in rat cardiac fibroblasts.

OBJECTIVE: To examine whether 17-beta-oestradiol (E(2)) may alter angiotensin II (Ang II) induced cell proliferation and to identify the putative underlying signalling pathways in rat cardiac fibroblasts. DESIGN: Cultured rat cardiac fibroblasts were preincubated with E(2) then stimulated with Ang II. [(3)H]Thymidine incorporation and endothelin-1 (ET-1) gene expression were examined. The effect of E(2) on Ang II induced NADPH oxidase activity, reactive oxygen species (ROS) formation, and extracellular signal regulated kinase (ERK) phosphorylation were tested to elucidate the intracellular mechanism of E(2) in proliferation and ET-1 gene expression. RESULTS: Ang II increased DNA synthesis, which was inhibited with E(2) (1-100 nmol/l). E(2), but not 17-alpha-oestradiol, inhibited Ang II induced ET-1 gene expression as shown by northern blotting and promoter activity assay. This effect was prevented by co-incubation with the oestrogen receptor antagonist ICI 182,780 (1 micromol/l). E(2) also inhibited Ang II increased NADPH oxidase activity, ROS formation, ERK phosphorylation, and activator protein-1 mediated reporter activity. CONCLUSIONS: The results suggest that E(2) inhibits Ang II induced cell proliferation and ET-1 gene expression, partially by interfering with the ERK pathway through attenuation of ROS generation. Thus, this study provides important new insight regarding the molecular pathways that may contribute to the proposed beneficial effects of oestrogen on the cardiovascular system.

Immediate and long-term outcomes of stent implantation for unprotected left main coronary artery disease.

Left main coronary artery (LMCA) disease is now uniformly treated with coronary artery bypass grafting (CABG). However, some patients with LMCA disease do not receive CABG because of high operative risks. The advent of stent implantation has permitted a non-operative improvement in myocardial blood flow in many patients with single- and multi-vessel coronary artery disease. However, the outcomes of stent implantation for unprotected LMCA disease are still unclear. Stent implantation was performed for unprotected LMCA disease in 13 patients; eight patients had high operative risk and five patients had refused CABG. The primary success rate was 100% (13/13 patients). One patient (8%) developed a non-Q-wave myocardial infarction after LMCA stenting. Repeat angiography was obtained in five patients (38%) with recurrent angina, and three patients (23%) received repeated percutaneous transluminal coronary angioplasty (PTCA) for LMCA restenosis. In the follow-up period of 18+/-3 months, 12 patients (92%) remained in satisfactory condition with no further need for surgical intervention. One patient (8%) ultimately required CABG, and she died after CABG at 3 months after LMCA stenting. In conclusion, although CABG remains the standard treatment for LMCA disease, the present study demonstrates that stent implantation is a safe and clinically beneficial revascularization procedure for unprotected LMCA disease in patients who have high operative risk as well as those who refuse CABG.

Adenovirus-mediated heme oxygenase-1 gene transfer inhibits the development of atherosclerosis in apolipoprotein E-deficient mice.

BACKGROUND: Increasing evidence supports the role of heme oxygenase-1 (HO-1) in cytoprotective response and iron homeostasis. The object of this study was to investigate whether adenovirus-mediated gene transfer of HO-1 in arteries reduces iron overload and inhibits lesion formation in apolipoprotein E (apoE)-deficient mice. METHODS AND RESULTS: Infection of rat aortic smooth muscle cells with adenovirus carrying the human HO-1 gene (Adv-HO-1) resulted in a high-level expression of HO-1 protein, which effectively reduced the hemin-induced iron overload in these cells. Adenovirus-mediated gene transfer in arteries in vivo was achieved by direct injection of Adv-HO-1 into the left ventricles of anesthetized animals. Transgene was expressed in the endothelium and aortic lesion of apoE-deficient mice after they had received recombinant adenovirus for 1 week and gradually decayed during the next 5 weeks. When young apoE-deficient mice (14 weeks old) received Adv-HO-1 (2.5 x 10(9) pfu) for 6 weeks, lesions that developed in the aortic root or aortic arch were significantly smaller than those in control littermates receiving empty viral vector. Furthermore, the iron deposition as well as tissue iron content was much less in aortic tissue of Adv-HO-1-treated mice. The inhibitory effect of HO-1 gene transfer on the progression of advanced lesions was also observed in older apoE-deficient mice (20 weeks old) receiving Adv-HO-1 intraventricularly. CONCLUSIONS: Overexpression of HO-1 in vascular cells facilitates iron metabolism and attenuates development of atherosclerosis in apoE-deficient mice.

Colocalization and interaction of cyclooxygenase-2 with caveolin-1 in human fibroblasts.

Results from our previous study suggest that cyclooxygenase-2 (COX-2) induced by phorbol 12-myristate 13-acetate (PMA) may be localized to caveolae-like structures (Liou, J.-Y., Shyue, S.-K., Tsai, M.-J., Chung, C.-L., Chu, K.-Y., and Wu, K. K. (2000) J. Biol. Chem. 275, 15314-15320). In this study, we determined subcellular localization of COX-2 and caveolin-1 by confocal microscopy. COX-2 in human foreskin fibroblasts stimulated by PMA (100 nm) or interleukin-1beta (1 ng/ml) for 6 h was localized to plasma membrane in addition to endoplasmic reticulum and nuclear envelope. Caveolin-1 was localized to plasma membrane, and image overlay showed colocalization of COX-2 with caveolin-1. This was confirmed by the presence of COX-2 and caveolin-1 in the detergent-insoluble membrane fraction of cells stimulated by PMA. Immunoprecipitation showed complex formation of COX-2 with caveolin-1 in a time-dependent manner. A larger quantity of COX-2 was complexed with caveolin-1 in PMA-treated than in interleukin-1beta-treated cells. Purified COX-2 complexed with glutathione S-transferase-fused caveolin-1, which was not inhibited by the scaffolding domain peptide. Caveolin-1-bound COX-2 was catalytically active, and its activity was not inhibited by the scaffolding domain peptide. These results suggest that COX-2 induced by PMA and interleukin-1beta is colocalized with caveolin-1 in the segregated caveolae compartment. Because caveolae are rich in signaling molecules, this COX-2 compartment may play an important role in diverse pathophysiological processes.

Selective augmentation of prostacyclin production by combined prostacyclin synthase and cyclooxygenase-1 gene transfer.

BACKGROUND: We tested the hypothesis that combined cyclooxygenase-1 (COX-1) and prostacyclin synthase (PGIS) gene transfer selectively augments prostacyclin production without a concurrent overproduction of other prostanoids. METHODS AND RESULTS: ECV304 cells were transfected with bicistronic pCOX-1/PGIS versus pCOX-1 or pPGIS, and prostanoids were analyzed. Contrary to the high prostaglandin E2 synthesis in pCOX-1 transfected cells, selective prostacyclin formation was noted with bicistronic plasmid transfection. Next, we determined the optimal ratio of Ad-COX-1 to Ad-PGIS by transfecting human umbilical vein endothelial cells with various titers of these 2 adenoviral constructs and determined the level of protein expression and prostanoid synthesis. Our results show that optimal ratios of adenoviral titers to achieve a large prostacyclin augmentation without overproduction of prostaglandin E2 or F2alpha were 50 to 100 plaque forming units (pfu) of Ad-COX-1 to 50 pfu of Ad-PGIS per cell. A higher Ad-PGIS to Ad-COX-1 ratio caused a paradoxical decline in prostacyclin synthesis. CONCLUSIONS: Prostacyclin synthesis can be selectively augmented by cotransfecting endothelial cells with an optimal ratio of COX-1 to PGIS. Combined COX-1 and PGIS gene transfer has the potential for therapeutic augmentation of prostacyclin.

Colocalization of prostacyclin synthase with prostaglandin H synthase-1 (PGHS-1) but not phorbol ester-induced PGHS-2 in cultured endothelial cells.

The subcellular colocalization of prostacyclin synthase (PGIS) with prostaglandin H synthase (PGHS) has not been delineated. To test the hypothesis that its colocalization with PGHS is crucial for prostacyclin synthesis, we determined subcellular locations of PGIS, PGHS-1, and PGHS-2 in bovine aortic endothelial cells by immunofluorescent confocal microscopy. PGIS and PGHS-1 were colocalized to nuclear envelope (NE) and endoplasmic reticulum (ER) in resting and adenovirus-infected bovine aortic endothelial cells. PGIS and PGHS-2 were also colocalized to ER in serum-treated or adenovirus-cyclooxygenase-2-infected cells. By contrast, PGIS was not colocalized with PGHS-2 in cells induced with phorbol 12-myristate 13-acetate where PGHS-2 was visualized primarily in vesicle-like structures. The lack of colocalization was accompanied by failed prostacyclin production. Resting ECV304 cells did not produce prostacyclin and had no detectable PGHS-1 and PGIS proteins. Confocal analysis showed abnormal colocalization of PGIS and PGHS-1 to a filamentous structure. Interestingly, the abundant PGIS and PGHS-1 expressed in adenovirus-infected ECV304 cells were colocalized to NE and ER, which synthesized a large quantity of prostacyclin. These findings underscore the importance of colocalization of PGHS and PGIS to ER and NE in prostacyclin synthesis.

Inhibition of octopus glutathione transferase by Meisenheimer complex analog, S-(2,4,6-trinitrophenyl) glutathione.

The tight binding of Meisenheimer intermediate with octopus digestive gland glutathione transferase was analyzed with 1,3,5-trinitrobenzene, which forms a trapped Meisenheimer complex with glutathione because there is no leaving group at the ipso carbon. By steady-state enzyme kinetic analysis, an inhibition constant of 1.89 +/- 0.17 microM was found for the transient formed, S-(2,4,6-trinitrophenyl) glutathione. The above inhibition constant is 407-fold smaller than the Km value for the substrate (2,4-dinitrochlorobenzene). Thus, S-(2,4,6-trinitrophenyl) glutathione is considered to be a transition-state analog. The tight binding of this inhibitor to the enzyme provides an explanation for the involvement of the biological binding effect on the rate enhancement in the glutathione transferase-catalyzed SNAr mechanism.

Effects of right ventricular pacing on ventriculoatrial conduction and systemic venous responses in sick sinus patients.

This study was designed to assess the ventriculoatrial (VA) conduction and systemic venous responses to right ventricular pacing at different pacing rates and the feasibility of identifying patients prone to pacemaker syndrome by means of a Doppler and two-dimensional echocardiographic technique. Twenty-two sick sinus patients who received ventricular-demand permanent pacemakers constituted the study group. The proximal inferior vena cava (IVC) diameters were measured by two-dimensional echocardiography. Fourteen patients had VA conduction by preimplant electrophysiologic study or paced electrocardiogram (Group II), while the other 8 patients presented no VA conduction (Group I). Abnormal systolic retrograde flow in the hepatic vein following each paced beat could be demonstrated in those patients with VA conduction in the basal state. In the 8 patients without VA conduction, the IVC diameters were significantly increased during rapid right ventricular pacing in those with left ventricular dysfunction (n = 4) as compared with those with normal left ventricular function (n = 4) (% increment at 120 beats per minute.

Transvenous permanent left ventricular pacing. A case report.

This case report presents a patient who inadvertently received transvenous permanent left ventricular pacing through an unexpected atrial septal defect. This lead malpositioning was proved by two-dimensional and transesophageal echocardiography. The abnormal pattern of electric activation was demonstrated by radionuclide phase image analysis. He has been followed up for a total of forty-three months with antiplatelet therapy and has been free from systemic embolic phenomena. A simple and readily available method that could lead to early recognition of lead malpositioning is reiterated and the various causes, methods of detection, and prognosis of left ventricular pacing are discussed.

Delayed localized right atrial tamponade by intrapericardial hematoma after aortic valve replacement: a case report.

Cardiac tamponade, early or late after cardiac surgery, is an uncommon while a potentially lethal condition in which the classic signs of tamponade may be absent. High index of suspicion and rapid diagnosis are mandatory for life-saving decompressive therapy. We herein reported a case of delayed localized right atrial tamponade caused by loculated intrapericardial hematoma 26 days after aortic valve replacement, which was recognized immediately by transthoracic two-dimensional echocardiography. The clinical status improved dramatically after surgical removal of the hematoma.

Usefulness of transesophageal echocardiography for the detection of left atrial thrombi in patients with rheumatic heart disease.

Transesophageal (TEE) and transthoracic (TTE) echocardiograms were performed in 110 patients with rheumatic heart disease to evaluate the usefulness of these methods for the detection of left atrial thrombi. TEE was better than TTE for detecting left atrial thrombi (21 vs 9). The thrombi not detected by TTE were in the left atrial appendage in ten and over the left atrial posterior wall in two. Patients with left atrial thrombi had significantly smaller mitral valve area (P less than 0.01) and greater left atrial dimension (P less than 0.05) than those without. All patients with left atrial thrombi had atrial fibrillation. Thirty-one patients underwent surgical intervention and 13 were found to have left atrial thrombi. TEE detected left atrial thrombi in all 13 patients with a sensitivity of 100%, specificity of 100%, and accuracy of 100%, while TTE detected left atrial thrombi in only nine of these 13 patients with a sensitivity of 69.2%, specificity of 100%, and accuracy of 87.1%. Thus, TEE is superior to TTE for the detection of left atrial thrombi, especially for those thrombi located in the left atrial appendage and along the left atrial posterior wall.

Transesophageal color Doppler flow mapping of iatrogenic left-to-right interatrial shunting after percutaneous transluminal mitral valvotomy.

Characteristics of transesophageal color Doppler flow mapping of iatrogenic left-to-right interatrial shunts were assessed in 58 patients, 1 to 994 days after percutaneous transluminal mitral valvotomy. Transesophageal color Doppler flow mapping detected 22 cases of interatrial shunt whereas transthoracic two-dimensional echocardiography visualized only five interatrial septal defects. Five types of color Doppler flow patterns of interatrial shunts were found: type 1, a bluish jet passing through the interatrial septum into the right atrium with a small bluish proximal flow in the left atrium (50%); type 2, a bluish jet passing through the interatrial septum into the right atrium without a proximal flow (13.6%); type 3, a predominant bluish proximal flow in the left atrium passing through the interatrial septum with minimal flow entering into the right atrium (18.2%); type 4, an "en face" bluish jet in the right atrium (4.5%); and type 5, a "wall jet" with proximal flow adhering to and entering into the interatrial septum (13.6%). Oximetry demonstrated increased pulmonary-to-systemic flow ratio (range 1.07 to 3.32) in 11 patients (50%), which was significantly correlated with the maximal jet area derived from color Doppler flow mapping (r = 0.80, P = 0.001). Thus, transesophageal color Doppler flow mapping is useful in detection of left-to-right interatrial shunts after percutaneous transluminal mitral valvotomy, and recognition of the variable types of color flow mapping may further help identify these atypical interatrial shunts.

Early detection of cerebral dysfunction by quantitative EEG during cardiopulmonary-bypass period for open-heart surgery.

Neuropsychiatric complications are well known risks of open heart surgery. In this regard cerebral damage is caused either by microembolism or insult from low perfusion pressure as a consequence of extracorporeal circulation. It seems probable that monitoring of the quantitative electroencephalography (QEEG) during the surgical procedure can help minimize the incidence of neurological sequelae through early detection. Moreover, it seems important to establish whether the pre-operative EEG has a predictive value for the possible neurological outcome and whether the post-operative EEG has conclusive value for the neurological outcome and, above all to ascertain whether the peri-operative QEEG contains information valuable on the diagnosis of final neurological outcome. To elucidate the feasibility of QEEG as a means to monitor the cerebral function a study was thus undertaken. Twenty-five ASA II-IV patients with age ranging from 25-79 y/o undergoing cardiac surgery under extracorporeal circulation were enrolled for study. The types of surgery varied from coronary artery by-pass graft (CABG), valvular replacement, and excision and grafting for aortic dissecting aneurysm. 16-channel EEGs were recorded with a Cadwell Spectrum 32 in accordance with the international 10-20 system. 48 epochs (1 epoch = 2.5 seconds) of EEG recordings of pre-bypass and post-bypass periods were measured and quantitatively analyzed. Pre- & post-operatively, psychoneurological assessments were performed and correlated with the QEEG. The parameters employed in the study were absolute power, relative power, power asymmetry, and coherence. All the results of two-session comparisons were transformed into z score values. Our preliminary study revealed that there exists a specific multivariate profile of QEEG for specific disease entity. And all the patients with z score less than or equal to 3.0 turned out to be free from any significant sequelate.

[Atrial tachycardia with block in congenital atrial septal defect. A case report].

Atrial arrhythmia is not uncommon in congenital atrial septal defect. Its incidence increases with age and the hemodynamic compromise of right ventricle. In the literature, atrial fibrillation is the most common presentation in middle-aged adults with congenital atrial septal defect, followed by atrial flutter and paroxysmal supraventricular tachycardia. However, atrial tachycardia with block has rarely been reported. We here report a case of 61-year-old male with congenital secundum atrial septal defect who presented atrial tachycardia with block without taking digitalis. Normal sinus rhythm restored after six-day use of Verapamil and Quinidine.

Misalignment characteristics of resonators formed by 90 degrees cone and mirror.

The stability condition of a resonator formed by a 90 degrees cone and mirror has been derived using ray tracing techniques. The result showed that this type of resonator is relatively insensitive to mirror misalignment. From this basis, the mirror misalignment tolerance was studied experimentally. Output energy and transverse mode characteristics vs. misalignment were measured, and the results show that this type of resonator does possess certain advantages over the two-mirror resonator.