Acute biological mitral valve thrombosis after the left atrial venting in a patient with a venoarterial extracorporeal membrane oxygenator.

BACKGROUND: Left atrial venting through atrial septotomy in patients with decreased left ventricular (LV) contractility after venoarterial extracorporeal membrane oxygenation (VA-ECMO) is a simple and effective method for treating LV decompression. MATERIALS & METHODS: We report a case of prosthetic mitral valve thrombosis after left atrial venting in a patient with VA-ECMO. RESULTS: In patients undergoing mitral valve replacement, left atrial venting reduces the flow through the mitral valve and forms a prosthetic thrombosis. DISCUSSION: Therefore, excessive left atrial venting should be avoided. Other venting methods that can maintain the flow through the mitral valve should be considered after mitral valve replacement.

Differentiation of multiple visceral artery aneurysm rupture from major bleeding after coronary artery bypass grafting.

BACKGROUND: Visceral artery aneurysm is a very rare disease, but it is clinically important because of the high risk of rupture involved. These ruptures must be differentiated from those that occur during hospitalization after extra-abdominal surgery. METHODS: During hospitalization after off-pump coronary artery bypass grafting, a 77-year-old woman developed hypovolemic symptoms and had decreased hemoglobin. There was no obvious bleeding, but while screening for possible complications after cardiac surgery, abdominal computed tomographic angiography showed multiple visceral artery aneurysms of the gastroduodenal and pancreaticoduodenal arteries along with hemoperitoneum. RESULTS: The patient underwent coil embolization of the visceral artery aneurysm and was discharged without any complications. CONCLUSIONS: In patients with coronary artery disease with risk factors for atherosclerosis, if anemia occurs without apparent bleeding after surgery, visceral artery aneurysm should be considered as a differential diagnosis.

Comparison of four facial muscles, orbicularis oculi, corrugator supercilii, masseter or mylohyoid, as best predictor of good conditions for intubation: a randomised blinded trial.

CONTEXT: Monitoring of facial muscles after neuromuscular blockade can give an early indication of respiratory muscle readiness for tracheal intubation. OBJECTIVE: To assess which facial muscle, the orbicularis oculi, corrugator supercilii, masseter or the mylohyoid, is the best predictor of readiness for intubation after rocuronium. DESIGN: Prospective, randomised, blinded trial. SETTING: Single centre: Seoul, Korea, from August 2012 to November 2012. PATIENTS: Two hundred and eighty-eight patients aged 22 to 64 years were randomised to one of eight study groups: orbicularis oculi, corrugator supercilii, masseter and mylohyoid for rocuronium 0.6 or 1.2 mg kg. INTERVENTION: The maximum twitch depression at the eyelid (orbicularis oculi), the superciliary arch (corrugator supercilii), the cheek (masseter) and the submental triangle (mylohyoid) was assessed after rocuronium 0.6 and 1.2 mg kg. Endotracheal intubation was performed after maximal neuromuscular blockade, and intubating conditions were appraised. MAIN OUTCOME MEASURES: The onset time of rocuronium and the quality of the intubation conditions were assessed. RESULTS: The onset times in the orbicularis oculi, corrugator supercilii and masseter were significantly faster than that in the mylohyoid (P < 0.001). 'Clinically acceptable' intubation conditions were significantly enhanced in the mylohyoid (94%) compared with those in the orbicularis oculi (80%) and masseter (78%) after rocuronium 0.6 mg kg (P < 0.05), and no difference with corrugator supercilii (92%). Despite differences in onset time of orbicularis oculi and masseter compared to mylohyoid (P < 0.05), intubating conditions were similar among the four muscles after rocuronium 1.2 mg kg. CONCLUSION: Following rocuronium 0.6 mg kg at similar depths of anaesthesia, the monitoring of the corrugator supercilii provided the best balance of a shorter onset time while maintaining 'clinically acceptable' intubation conditions. TRIAL REGISTRATION: IRB File No.: HYUH 2012-07-009.

Creation of immortalised epithelial cells from ovarian endometrioma.

BACKGROUND: Epithelial cells of endometriotic tissues are difficult to propagate in vitro as experimental material is scarce owing to their limited life span. However, there is an increasing concern regarding their malignant transformation in ovaries. The present study sought to generate their stable culture system. METHODS AND RESULTS: Purified epithelial cells isolated from ovarian endometriomas using microscopic manipulation were successfully immortalised by combinatorial transfection of human cyclinD1, cdk4 and human telomerase reverse transcriptase (hTERT) genes, whereas the introduction of hTERT alone, or together with cdk4, was insufficient for immortalisation, leading to cellular senescence. We confirmed stable cytokeratin expression in the immortalised cells, proving their epithelial origin. These cells expressed progesterone receptor B and showed significant growth inhibition by various progestins. Oestrogen receptor (ER) expression was detected in these cells, albeit at low levels. Additional overexpression of ERα generated stable cells with oestrogen-dependent growth activation. Soft-agar colony formation assay and nude mice xenograft experiments demonstrated that these cells, even those with additional inactivation of p53, did not have transformed phenotypes. CONCLUSION: We for the first time generated immortalised epithelial cells from ovarian endometrioma that retained sex steroid responsiveness. These cells are invaluable tools not only for the consistent in vitro work but also for the study of molecular pathogenesis or carcinogenesis of endometriosis.

Circulating tumour cells detected by a novel adenovirus-mediated system may be a potent therapeutic marker in gynaecological cancers.

BACKGROUND: Recently developed detection system for circulating tumour cells (CTCs) using a telomerase-specific replicative adenovirus generated nonspecific green fluorescent protein (GFP) signals because of the co-presence of white blood cells (WBCs) nonspecifically infected by viruses. Here, we established a unique detection system for CTCs that completely excludes nonspecific signals. METHODS: Blood obtained from the patients was subjected to haemolytic processes to eliminate red blood cells. The cell pellets were then infected with OBP-401, fixed, incubated with fluorescence-labelled anti-CD45 antibody to mark white blood WBCs, and examined on slides under a microscope. RESULTS: Preparatory experiments with cancer cells artificially added to healthy donor samples confirmed that CD45 labelling could distinguish GFP-positive cancer cells from WBCs. In 53 patients with gynaecological cancers, CTCs were detected in 21 patients (39.6%) when CD45-positive cells were excluded as WBCs among GFP-positive cells. No CTCs were detected in samples from healthy volunteers. There was no significant correlation between CTC counts and known clinicopathological factors. The CTCs rapidly vanished after surgery or chemotherapy in most patients whose treatments were effective. In contrast, the persistence of CTCs even after treatments was tightly associated with poor response to the treatments (P<0.005). CONCLUSION: The presence of CTCs in our system may potentially be a novel therapeutic marker in gynaecological cancers.

The efficacy of several neuromuscular monitoring modes at the P6 acupuncture point in preventing postoperative nausea and vomiting.

BACKGROUND: In this study, we tested the efficacy of several neuromuscular monitoring modes at the P6 acupuncture point for preventing postoperative nausea and vomiting (PONV). METHODS: In this prospective, double-blind, randomized, placebo-controlled trial, 264 women undergoing laparoscopic hysterectomy were evaluated for PONV. Neuromuscular blockade was monitored by acceleromyography with 1-Hz single twitch (ST) over the ulnar nerve (n = 54, control), and ST (n = 52), train-of-four (n = 53), double-burst stimulation (n = 53), or tetanus (n = 52) over the median nerve stimulating at the P6 acupuncture point. RESULTS: The incidence of PONV (P = 0.022), the number of requests for patient-controlled analgesia (P = 0.009), and total patient-controlled analgesia volume (P = 0.042) 6 hours after tetanic stimulation were significantly reduced in the treatment group compared with the control group. Overall, patients in the tetanus group were more satisfied with the management of PONV compared with patients in the control group. CONCLUSION: Tetanic stimulation applied to the P6 acupuncture point can reduce PONV after laparoscopic hysterectomy compared with ST stimulation of the ulnar nerve, resulting in a greater degree of patient satisfaction. None of the stimulations, ST, train-of-four, or double-burst, applied to the P6 acupuncture point significantly affected PONV.

Expression of HER-2 affects patient survival and paclitaxel sensitivity in endometrial cancer.

BACKGROUND: Disabled phosphatidylinositol 3-kinase (PI3K)/AKT and mitogen-activated protein kinase/extracellular signal-regulated kinase signalling is involved in endometrial carcinogenesis, and there is evidence that expression of epidermal growth factor receptor (EGFR) family members has a role in such intracellular signalling pathways. This study analysed the prognostic impact of EGFR family expression in endometrial cancer in relation to PI3K-AKT and MAPK-ERK signalling, as well as drug sensitivity. METHODS AND RESULTS: Immunohistochemical analysis using 63 surgical specimens of endometrioid-type endometrial cancers revealed that EGFR, human epidermal growth factor receptor (HER)-2 and HER-4 were expressed in 25 (39.7%) of 63, 26 (41.3%) of 63 and 31 (49.2%) of 63 tumours, respectively. Gene amplification of HER-2 was observed in 2 of 26 patients with high HER-2 expression. Kaplan-Meier analysis revealed that high HER-2 expression was a factor that negatively influenced the progression-free and overall survival rate (P<0.05), and multivariate analysis showed high HER-2 expression to be an independent prognostic factor. Subsequently, we performed in vitro knockdown analysis to investigate the linkage between HER-2 expression and PI3K-AKT pathways. Short interfering RNA (siRNA)-based knockdown of HER-2 in endometrial cancer cells led to a significant reduction in phosphorylated AKT (p-AKT) expression, indicating the existence of a HER-2/PI3K-AKT axis. As the PI3K-AKT pathway is known to have crucial roles in anticancer drug sensitivity, we examined the involvement of HER-2 in sensitivity to paclitaxel. Short interfering RNA-based knockdown of HER-2 conferred increased sensitivity to paclitaxel in endometrial cancer cells, attenuating the induction of p-AKT on paclitaxel stimulation, which was cancelled by inactivating AKT by the introduction of a dominant-negative form. CONCLUSION: HER-2 is a significant prognostic factor of endometrioid-type endometrial cancer, as well as a key molecule that affects paclitaxel sensitivity by HER-2 interaction with the PI3K-AKT pathway.

Krüppel-like factor 4 mediates anti-proliferative effects of progesterone with G0/G1 arrest in human endometrial epithelial cells.

Activation of the progesterone receptor (PR) inhibits cell proliferation in various reproductive tissues. However, the molecular mechanisms underlying the regulation of cell proliferation by PR remain poorly understood. It is well established that Krüppel-like factor 4 (KLF4), a family of zinc fingercontaining transcription factors, induces cell cycle arrest in epithelial cells. In this study, we investigated whether KLF4 served as a target of PR activation during cell proliferation using human endometrial epithelial cells. PR agonists, progesterone and dienogest, were found to produce a lasting increase in the expression of KLF4 mRNA, followed by a decrease in cyclin D1 mRNA, and inhibit cell proliferation with G0/G1 arrest. KLF4 knockdown using KLF4 small interferingRNA abrogated the inhibition of cell proliferation by PR agonists. In addition, forced expression of KLF4 inhibited cyclin D1 promoter transactivation. These results suggest that PR agonists induce KLF4 expression and then inhibit cyclin D1 expression, and consequently inhibit cell proliferation in human endometrial epithelial cells. In terms of human reproductive tissue, KLF4 may be a factor concerning cell cycle, directly responsive to PR activation.

Capsicum plaster at the Hegu point reduces postoperative analgesic requirement after orthognathic surgery.

BACKGROUND: Capsicum plaster at classical Chinese acupoints is an alternative to acupuncture, which has been used as an effective method for preventing postoperative nausea and vomiting, sore throat, and pain. In this study, we investigated the postoperative analgesic efficacy of capsicum plaster at Hegu (LI 4) acupoints in patients after bilateral sagittal split ramus osteotomy. METHODS: A double-blind, sham-controlled study was conducted with 84 patients undergoing orthognathic surgery, and who were randomly assigned to three treatment regimens (n = 28 each): Hegu group = capsicum plaster at Hegu acupoints and placebo tape on the shoulders as a nonacupoint; sham group = capsicum plaster on the shoulders and placebo tape at Hegu acupoints; and control group = placebo tape at Hegu acupoints and on the shoulders. The capsicum plaster was applied before induction of anesthesia and maintained for 8 h per day for 3 postoperative days. RESULTS: The total amount of patient-controlled analgesia, containing 6.5 microg/mL fentanyl and 1.2 mg/mL ketorolac, administered in the first 24 h after the operation was decreased in the Hegu group (26.8 +/- 3.4 mL) compared with the control (44.2 +/- 7.3 mL) and sham (42.1 +/- 6.9 mL) groups (P < 0.01). The incidence of postoperative nausea and vomiting and the need for rescue medication were reduced, and the overall satisfaction score was greater in the Hegu group compared with other groups (P < 0.01). CONCLUSION: The capsicum plaster at the Hegu acupoints decreased the postoperative opioid requirements and opioid-related side effects in patients after orthognathic surgery.

Creation of tumorigenic human endometrial epithelial cells with intact chromosomes by introducing defined genetic elements.

Several genetic mutations have been identified in human endometrial cancers, but the specific combinations of mutations required to form endometrial cancer cells remain unknown. In the present study, we established an in vitro model of endometrial carcinogenesis, in which defined genetic elements were introduced into endometrial epithelial cells to create transformed endometrial cells at different stages. Introduction of the human papillomavirus type 16 E6/E7 gene and the human telomerase reverse transcriptase (hTERT) gene into human primary endometrial epithelial cells was sufficient to generate immortalized cells. Introduction of hTERT in early passages stabilized telomeres and created immortalized cells with normal karyotype, whereas introduction of hTERT in later passages generated immortalized cells but with widespread chromosome abnormalities. However, neither of those two immortalized cell lines exhibited tumorigenic phenotypes. Tumorigenic endometrial epithelial cells with invasive capacity were created by introducing a mutant K-ras allele into immortalized cells, keeping their chromosomes intact. Inhibiting the PTEN gene and activating Akt pathways did not create tumorigenic phenotypes, although the latter conferred anchorage-independent growth capacity. These findings suggest that neoplastic transformation of human endometrial cells can occur in the absence of widespread chromosomal abnormality, and that the combination of Rb inactivation, telomerase activation and altered K-ras signaling is sufficient for in vitro neoplastic transformation. The present experimental model can help clarify the genetic requirements for endometrial carcinogenesis, and it is useful for testing and developing specific inhibitors of specific oncogenic pathways.

Early growth response-1 mediates up-regulation of telomerase in placenta.

Telomerase is thought to play a very important role in oncogenesis. It is also believed to wind back the "mitotic clock" which leads to ageing and enable permanent cell division. We evaluated telomerase activity in chorionic tissues, with particular attention to the early growth response-1 (EGR-1) gene, the importance of what was recently shown by Khachigian et al. We started our study by evaluating the relationship between activation of transcription of the human telomerase reverse transcriptase (hTERT) gene and EGR-1 gene. For this purpose, we first evaluated telomerase activity using the villous cancer cell lines JAR and JEG-3. We then demonstrated that EGR-1 plays an important role in activation of the transcription of hTERT by luciferase assay using hTERT promoter constructs. As a result of further computer analysis, we discovered a site postulated to be an EGR-1 consensus binding site at -273 to -281 in the hTERT promoter region. With forced expression of EGR-1, an increase in hTERT protein concentration was detected on Western blot analysis, while marked high expression of hTERT mRNA was observed by reverse transcriptase polymerase chain reaction. Furthermore, we evaluated the expression of EGR-1 and hTERT at the mRNA level in the placenta during the first, second and third trimesters of pregnancy and in patients with preeclampsia. Expression of EGR-1 and hTERT in the chorion increased in the first trimester of pregnancy and decreased later. Increased expression was noted in the placenta of patients with preeclampsia. The present findings suggest that EGR-1 plays an important role in activating the transcription of hTERT, showing that activation of the transcription of hTERT by EGR-1 is involved in the trophoblast growth mechanism.

Analysis of outcome of Stage I-III endometrial cancer treated with systematic operation omitting paraaortic lymphadenectomy.

PURPOSE: The aim of this study was to assess the outcomes of endometrial cancer patients treated with systematic surgery omitting paraarotic lymphadenectomy. PATIENTS AND METHODS: We retrospectively analyzed a consecutive series of 84 endometrioid-type endometrial cancer patients at FIGO Stage I, II or III without grossly metastatic paraaortic lymphadenodes, who underwent surgery at our institute. RESULTS: Sixty-five patients (77%) underwent primary surgery with pelvic lymphadenectomy while the remaining 19 patients underwent surgery without lymphadenectomy due to severe medical complications or age greater than 70 years. The patients with high risk for recurrence were treated mainly by adjuvant irradiation therapy of the whole pelvis. The median follow-up period was 44 months. The 5-year overall survival (OS) rate was 92%, 92% and 65% for FIGO Stage I, II and III, respectively. Recurrence was detected in eight of the 82 optimally operated patients (9.8%). Out of the eight recurrent patients, five patients had a recurrent tumor at extra-pelvic sites (chest or abdomen), two patients had a recurrent tumor only in a paraaortic lymph node, and one patient had a recurrent tumor only in the vagina. Thus, the recurrence rate was relatively low, with 2.4% relapse at the paraarotic lymph nodes, and 5-year OS rate appeared to be favorable. However, all the six recurrent patients who underwent adjuvant radiation therapy had distant recurrence. CONCLUSIONS: These findings indicate that omission of paraarotic lymphadenectomy may be acceptable for endometrial cancer patients without gross metastasis at this site. However, the high rate of distant recurrence after whole pelvic irradiation strongly indicates an urgent need to develop potent systemic adjuvant therapy, potentially by chemotherapy or chemoradiation therapy.

[Ventricular septal perforation due to weak blunt chest trauma; report of a case].

We report a 67-year-old female patient with ventricular septal perforation after weak blunt chest trauma. She tumbled down on a frozen street. Approximately 1 week later, the patient was aware of shortness of breath on exertion. On admission, holosystolic murmur was detected on chest wall and routine electrocardiogram examination showed ST-T change which suggested myocardial ischemia. Acute myocardial infarction and ventricular septal defect with left-to-right shunt was suspected. The echocardiography and cardiac catheterization revealed the muscular type ventricular septal perforation near the apex with large left-to-right shunt flow (82% shunt ratio). The congestive heart failure was controlled successfully by conservative medical treatment. Surgical repair was scheduled on the 28th day after initial chest trauma because of large left-to-right shunt. A hole of about a diameter of 2 cm with fibrous edge of the muscular septum was closed through a left ventriculotomy using a Dacron patch under cardiopulmonary bypass. Postoperative course was uneventful and the patient was discharged without symptoms of congestive heart failure.

[Coronary malperfusion due to acute type A aortic dissection; surgical strategy and results].

Coronary malperfusion due to acute type A aortic dissection (DAA) is a lethal complication. It is especially difficult to rescue the patients with left coronary malperfusion because of acute global myocardial infarction (AMI), even with successful surgical treatments, including the replacement of the ascending aorta and coronary artery bypass grafting (CABG). We review our experience and illustrate our approach to these critically ill patients. In addition, we classify the mechanism of malperfusion into 4 types based upon perioperative findings and discuss surgical management indivisually. From January 1990 to April 2005, a total of 260 patients were operated for DAA in our institution. Twenty (7.7%) patients, 11 men and 9 women were suffering from coronary malperfusion due to DAA. The mean age was 55 (range 28-72) years. The right coronary artery was involved in 9 patients, and the left in 11. All procedures such as graft replacement and CABG were done on an emergent or urgent basis. Hospital mortality rate of right coronary malperfusion was 22% (2/9 patients), and that related to left coronary malperfusion was 5/11 (45%). Assisting device was required in 9 cases, veno-arterial bypass (VAB) in 6 cases, left ventricular assist system (LVAS) in 1, left heart bypass (LHB) in 1, LHB+right heart bypass (RHB) in 1. We lost all patients using VAB. Only 3 patients supported with strong assist device survived. Aggressive myocardial resuscitation and early operation are the key factors in the management of these critically ill patients. But once severe myocardial infarction occurs, V-A bypass (percutaneous cardiopulmonary support) is useless in treating patients with DAA who develop severe heart failure. We recommend to implant stronger assist device including LVAS immediately before exacerbation of multiple organ failure. In conclusion, surgical management is not easy for emergency patients with DAA in association with myocardial ischemia. However, reasonable surgical results can be obtained with supplemental CABG and strong mechanical support of the left ventricle.

The analgesic effects of capsicum plaster at the Zusanli point after abdominal hysterectomy.

BACKGROUND: Acupuncture has been used to supplement opioid analgesics for postoperative pain control. We designed this double-blind, sham-controlled study to assess the effectiveness of capsicum plaster (PAS) at Zusanli (ST-36) acupoints on postoperative opioid analgesic requirement, side effects, and recovery profile. METHODS: Ninety women undergoing total abdominal hysterectomy were randomly assigned to 3 treatment regimens (n = 30 each): group Zusanli = PAS at Zusanli acupoints, group sham = PAS at the nonacupoints on the shoulders, and group control = placebo tape at Zusanli acupoints. The PAS was applied before induction of anesthesia and maintained for 8 h per day for 3 postoperative days. RESULTS: The total amount of morphine administered in the first 24 h after the operation was significantly decreased in group Zusanli (31.5 +/- 6.8 mL) compared with groups control (44.3 +/- 10.1 mL) and sham (44.6 +/- 10.4 mL) (P < 0.01). The incidence of postoperative side effects and the use of rescue antiemetics during the 72 h after surgery were significantly reduced in group Zusanli compared with other groups (P < 0.01). CONCLUSION: PAS at Zusanli points decreased the postoperative opioid requirement and opioid-related side effects of patients undergoing abdominal hysterectomy.

E2F-1 represses transcription of the human telomerase reverse transcriptase gene.

The ends of human chromosomes (telomeres) lose up to 200 bp of DNA per cell division. Chromosomal shortening ultimately leads to senescence and death in normal cells. Many human carcinoma lines are immortal in vitro, suggesting that these cells have a mechanism for maintaining the ends of their chromosomes. Telomerase is a ribonucleoprotein complex that synthesizes telomeric DNA onto chromosomes using its RNA component as template. Telomerase activity is found in most tumor cells, but is absent from normal cells. Little is known about how normal human cells repress telomerase (hTERT) gene expression. Mice carrying an E2F-1 null mutation develop a variety of malignant tumors, suggesting that this transcription factor has a tumor suppressor function. To determine mechanisms by which E2F-1 suppresses tumor formation, we examined the role of this transcription factor in regulation of the hTERT promoter in human cells. We identified two putative E2F-1-binding sites proximal to the transcriptional start site of the hTERT promoter. Mutation of these sites produced dramatic increases in promoter activity. Overexpression of E2F-1 but not a mutant E2F-1 repressed hTERT promoter activity in reporter gene assays. This repression was abolished by mutation of the E2F-1-binding sites in the hTERT promoter. Human cancer cell lines stably overexpressing E2F-1 exhibited decreased hTERT mRNA expression and telomerase activity. We conclude that E2F-1 has an atypical function as a transcriptional repressor of the hTERT gene in human cells.

Sp1 cooperates with c-Myc to activate transcription of the human telomerase reverse transcriptase gene (hTERT).

Telomerase activation is thought to be a critical step in cellular immortalization and carcinogenesis. The human telomerase catalytic subunit (hTERT) is a rate limiting determinant of the enzymatic activity of human telomerase. In the previous study, we identified the proximal 181 bp core promoter responsible for transcriptional activity of the hTERT gene. To identify the regulatory factors of transcription, transient expression assays were performed using hTERT promoter reporter plasmids. Serial deletion assays of the core promoter revealed that the 5'-region containing the E-box, which binds Myc/Max, as well as the 3'-region containing the GC-box, which binds Sp1, are essential for transactivation. The mutations introduced in the E-box or GC-box significantly decreased transcriptional activity of the promoter. Overexpression of Myc/Max or Sp1 led to significant activation of transcription in a cell type-specific manner, while Mad/Max introduction repressed it. However, the effects of Myc/Max on transactivation were marginal when Sp1 sites were mutated. Western blot analysis using various cell lines revealed a positive correlation between c-Myc and Sp1 expression and transcriptional activity of hTERT. Using fibroblast lineages in different stages of transformation, we found that c-Myc and Sp1 were induced to a dramatic extent when cells overcame replicative senescence and obtained immortal characteristics, in association with telomerase activation. These findings suggest that c-Myc and Sp1 cooperatively function as the major determinants of hTERT expression, and that the switching functions of Myc/Max and Mad/Max might also play roles in telomerase regulation.

Telomerase activation by histone deacetylase inhibitor in normal cells.

Although telomerase activity is known to be regulated mainly at the level of transcription of the human telomerase catalytic subunit (hTERT) gene, the molecular mechanism underlying tumor-specific expression of telomerase remains unclear. Emerging evidence suggests that reversible acetylation of nucleosomal histones and the resultant changes in the chromatin structure are important processes in gene transcription. In particular, histone deacetylase (HDAC) inhibitors activate the transcription of certain genes by altering the acetylation status of nucleosomal histones. The present study examines the effects of HDAC inhibitor on hTERT gene transcription. Treatment with tricostatin A (TSA) induced significant activation of hTERT mRNA expression and telomerase activity in normal cells, but not in cancer cells. Transient expression assays revealed that TSA activates the hTERT promoter. Furthermore, the proximal 181 bp core promoter of hTERT, which contains two c-Myc and five Sp1 sites, was determined to be the responsible element. Overexpression of Sp1 enhanced responsiveness to TSA, and mutation of Sp1 sites, but not c-Myc sites, of the core promoter of hTERT abrogated this activation. Introduction of the dominant-negative form of the Sp family inhibited TSA activation. These results indicate that HDAC inhibitor activates the hTERT promoter in normal cells, in which Sp1 plays a key role. This finding suggests one way whereby histone deacetylation may be involved in silencing the hTERT gene in normal cells.

Identification and characterization of negative regulatory elements of the human telomerase catalytic subunit (hTERT) gene promoter: possible role of MZF-2 in transcriptional repression of hTERT.

Human telomerase reverse transcriptase (hTERT) is a catalytic subunit of human telomerase and is a critical determinant of the enzymatic activity of telomerase. Expression of hTERT is known to be regulated mainly at the transcriptional level. In the present study, using transient expression assays, we identified a 400 bp silencer of the hTERT promoter between -776 and -378 upstream of the proximal core promoter. The inhibitory effects of this silencer were enhanced with cellular differentiation. A computer-assisted homology search identified multiple binding motifs for myeloid-specific zinc finger protein 2 (MZF-2) within this region. Mutation introduced in these sites resulted in significant activation of hTERT transcription. Gel shift assays demonstrated that MZF-2 proteins specifically bound to these sites. Overexpression of MZF-2 in cells led to down-regulation of hTERT transcription as well as telomerase activity. These findings suggest that the 400 bp region upstream of the hTERT core promoter that we identified functions as a negative regulatory region and that MZF-2 may be an effector of negative regulation of hTERT.

Prevention of withdrawal associated with the injection of rocuronium in adults and children.

STUDY OBJECTIVE: To determine which technique prevents the withdrawal associated with rocuronium administration in adults and children. DESIGN: Blinded, randomized, prospective trial. SETTING: This study was set at an inpatient anesthesia in a university teaching hospital. PATIENTS: 200 adult patients (aged 19-63 years) and 150 children (aged 2-9 years) undergoing elective surgery requiring endotracheal intubation. INTERVENTIONS: Four groups in adult and 3 groups in children of 50 patients each were investigated. In adult study, control groups with free intravenous (IV) flow (C-F) or the occlusion of IV flow (C-O) received saline as the pretreatment of rocuronium; lidocaine groups with free IV flow (L-F) or the occlusion of IV flow (L-O) received lidocaine as the pretreatment of rocuronium, preceded by thiopental 5 seconds before. In children study, groups P and L received saline and lidocaine as the pretreatment of rocuronium, respectively, and group S received rocuronium mixed with sodium bicarbonate after the pretreatment of placebo preceded by thiopental. MEASUREMENTS AND MAIN RESULTS: The patient's response to rocuronium injection was graded using a 4-point scale. The pH and osmolality of treatment solution were measured. The incidence of no movement after rocuronium was 96% in L-O, 46% in L-F, 26% in C-O, and 18% in C-F in adult and 96% in S, 58% in L, and 8% in P in children. CONCLUSIONS: Withdrawal after rocuronium can be eliminated by the pretreatment of lidocaine during the occlusion of the IV flow in adults and addition of sodium bicarbonate in children.