6-mercaptopurine (6-MP) induces p53-mediated apoptosis of neural progenitor cells in the developing fetal rodent brain.

6-mercaptopurine (6-MP), a DNA-damaging agent, induces apoptosis of neural progenitor cells, and causes malformation in the fetal brain. The aim of the present study is to clarify the molecular pathway of 6-MP-induced apoptosis of neural progenitor cells in the fetal telencephalon of rats and mice. p53 protein is activated by DNA damage and induces apoptosis through either the intrinsic pathway involving the mitochondria or the extrinsic pathway triggered by death receptors. In this study, the expression of puma and cleaved caspase-9 proteins, which are specific intrinsic pathway factors, increased in the rat telencephalon after 6-MP treatment. 6-MP-induced apoptosis of neural progenitor cells was completely absent in p53-deficient mice. On the other hand, the expression of Fas protein, an extrinsic pathway factor, did not change throughout the experimental period in the rat telencephalon treated with 6-MP. The number of apoptotic neural progenitor cells was similar among Fas-mutated lpr/lpr and wild-type mice, suggesting that the Fas pathway does not play a significant role in 6-MP-induced apoptosis of neural progenitor cells. These results may suggest that the p53-mediated intrinsic pathway is essential for 6-MP-induced apoptosis of neural progenitor cells in the developing telencephalon of rats and mice.

Time-course changes in neural cell apoptosis in the rat fetal brain from dams treated with 6-mercaptopurine (6-MP).

6-Mercaptopurine (6-MP), one of the major drugs for the therapy of acute lymphoblastic leukemia and autoimmune diseases, is incorporated as thioguanine in nucleic acid and it induces cytotoxicity and fetotoxicity. In the present study, pregnant rats were treated with 50 mg/kg of 6-MP on 13 embryonic days (E), and fetuses were collected from 12 to 96 h after the treatment to examine the mechanism and time-course changes in neural cell death in the developing brain. The weights of fetal telencephalon and the thickness of the dorsal telencephalic wall of the fetuses were significantly reduced at 96 h. The number of pyknotic neural cells in the fetal telencephalon began to increase at 24 h, peaked at 36 h, and then gradually decreased toward 72 h. The nuclei of most of these pyknotic cells were stained positively by TUNEL method, which detects DNA fragmentation. Moreover, pyknotic cells were immunohistochemically positive for cleaved caspase-3, one of the key executioners of apoptosis, and the increased expression of the protein from 30 to 48 h was confirmed by using Western blot analysis. Also, electron microscopical features of the pyknotic cells showed ultrastructural characteristics of apoptosis. On the other hand, the number of mitotic and BrdU-positive neural cells in the telencephalon decreased from 30 to 72 h. These results suggest that 6-MP induced apoptotic cell death in neural cells in the rat fetal brain is probably due to cytotoxic action of 6-MP.

6-Mercaptopurine (6-MP) induces cell cycle arrest and apoptosis of neural progenitor cells in the developing fetal rat brain.

6-Mercaptopurine (6-MP), an analogue of hypoxanthine, is used in the therapy of acute lymphoblastic leukemia and causes fetal neurotoxicity. To clarify the mechanisms of 6-MP-induced fetal neurotoxicity leading to the cell cycle arrest and apoptosis of neural progenitor cells, pregnant rats were treated with 50 mg/kg 6-MP on embryonic day (E) 13, and the fetal telencephalons were examined at 12 to 72 h (h) after treatment. Flow-cytometric analysis confirmed an accumulation of cells at G2/M, S, and sub-G1 (apoptotic cells) phases from 24 to 72 h. The number of phosphorylated histone H3-positive cells (mitotic cells) decreased from 36 to 72 h, and the phosphorylated (active) form of p53 protein, which is a mediator of apoptosis and cell cycle arrest, increased from 24 to 48 h. An executor of p53-mediated cell cycle arrest, p21, showed intense overexpression at both the mRNA and protein levels from 24 to 72 h. Cdc25A protein, which is needed for the progression of S phase, decreased at 36 and 48 h. In addition, phosphorylated cdc2 protein, which is an inactive form of cdc2 necessary for G2/M progression, increased from 24 to 48 h. These results suggest that 6-MP induced G2/M arrest, delayed S-phase progression, and finally induced apoptosis of neural progenitor cells mediated by p53 in the fetal rat telencephalon.

[Surgical technique of aortic valve replacement for small aortic annulus in elderly patients].

Recent reports have shown that aortic valve replacement in elderly patients over 65 years with atherosclerotic aortic stenosis and a small aortic annulus is possible by using a small sized bioprosthesis (Carpentier-Edwards pericardial valve). Here we present out surgical technique. Firstly, the native calcified aortic valve was removed completely to gain total exposure of the surrounding aortic root and sinus of Valsalva like Bentall procedure. Secondly, a small sized bioprosthesis was implanted with intermittent noneverting mattress 2-0 sutures with spaghetti and small polytetrafluoroethylene (PTFE) felt. Aortic annulus is the dilated by inserting Hegar dilator sizing from 25 to 27 mm. Therefore, aortic valve replacement for small aortic annulus in intra- or supra-annular position should be easily accomplished. Good surgical results and hemodynamic state were achieved in 25 consecutive cases using this technique.

[Isolated coronary artery bypass grafting in octogenarians].

We evaluate the outcomes of isolated coronary artery bypass grafting (CABG) in high-risk octogenarians. There are 20 cases in 5 years, which contain 1 re-operation case, and 10 emergency cases. Nineteen cases were performed off-pump CABG (OPCAB), and conventional CABG was performed for only 1 case because of unstable hemodynamics with intraaortic balloon pumping (IABP). We generally performed CABG with aorta no-touch technique. There is no death case, and no serious complication like stroke. We used 23 artery and 11 vein grafts for CABG, and mean number of grafts per patient was 1.8+/-0.7. And "hybrid therapy" with percutaneous transluminal coronary angioplasty (PTCA) was done for 5 cases. Octogenarians are at increased risk for perioperative morbidity and mortality after CABG, but we have performed OPCAB for them and have had good results. And we think that aorta no-touch technique, proper selection and use of graft are important.

Left ventricle restoration in patients with non-ischemic dilated cardiomyopathy: risk factors and predictors of outcome and change of mid-term ventricular function.

OBJECTIVE: The partial left ventriculectomy (PLV) for end-stage dilated cardiomyopathy (DCM) which worked in some patients has been reported, although the hospital mortality is high. To reduce hospital mortality, we selected operative procedures of left ventricular (LV) restoration to improve the operative results. We analyzed the risk factors and predictors of outcome, and the mid-term changes of the LV function were determined. PATIENTS AND METHODS: Between December 1996 and September 2000, 74 patients with non-ischemic DCM received LV restoration. The age ranged from 14 to 76 years (mean, 49.0+/-14.0 years), and there were 63 men and 11 women. The etiology of the DCM was idiopathic DCM in 49 patients, and dilated hypertrophic cardiomyopathy in seven patients and others in 18. The preoperative New York Heart Association (NYHA) functional class was 29 in class III and 45 in class IV, in which 32 patients depended on inotropic support. PLV or septal anterior ventricular exclusion (SAVE) was selected depending on the akinetic lesion of the LV based on the intraoperative echo-test. Fifty-six patients received elective operations, and emergency operations were performed in 18 patients. The risk factors and predictors of outcome were analyzed in 74 patients, and in 35 patients who survived more than 1 year after receiving LV restoration, the mid-term cardiac function was examined by cardiac echogram and catheterization. RESULTS: PLV was performed in 62 patients and SAVE in 12 patients. Concomitant mitral surgery was performed in 66 patients (89%) and tricuspid annuloplasty in 42 patients (57%). There were 15 hospital deaths and 13 patients died after discharge from the hospital (cardiac deaths in nine and non-cardiac deaths in four). In the 46 late survivors, the NYHA class was I or II in 42 patients and III in four patients. Selection of the procedure of LV restoration (P<0.01), elective operation (P<0.05), and the preoperative volume of LV (endodiastolic volume index of <180 ml/m(2); P<0.05) were risk factors and predictors influencing hospital and late death. After the operation, the LV function improved significantly and the improvement was maintained at the mid-term period; the LV ejection fraction was 31.8+/-7.9% (P<0.01) at 1 year from 23.0+/-7.3% preoperatively, left ventricular diastolic diameter was 62.8+/-10.9 (P<0.01) from 81.7+/-8.2 mm and the LV endosystolic volume index was 88.5+/-45.8 (P<0.05) from 162.6+/-41.6 ml/m(2). CONCLUSIONS: The operative results improved with the selection of the procedures, with elective operation, and mitral plasty for less cardiac dilatation. The mid-term results of clinical status and LV function showed the effectiveness of the operation.

[Left ventriculoplasty for non-ischemic cardiomyopathy with severe heart failure in 70 patients].

OBJECTIVES: Treatment of cardiac failure due to non-ischemic cardiomyopathy by left ventriculoplasty using partial left ventriculectomy (Batista operation) or septal anterior ventricular exclusion was evaluated. METHODS: Left ventriculoplasty was performed in 70 patients (59 men and 11 women with a mean age of 51 years) from December 1996 to June 2000. Preoperative New York Heart Association (NYHA) functional class was IV in 43 patients including 29 receiving inotropic support, and class III in 27 patients. Nineteen patients required emergency surgery because of on-going shock and 51 patients were operated electively. Combined cardiac procedures were mitral valve reconstruction in 62 patients (45 replacements, 17 repairs), tricuspid annuloplasty in 37, and aortic valve replacement in 4. The initial 24 patients underwent typical Batista operation regardless of myocardial viability. The other 46 patients underwent selective ventriculoplasty to excise or exclude the weakest part according to the findings of the intraoperative echo-guided volume reduction test. RESULTS: The intraaortic balloon pump was used in 12 patients and the left ventricular assist device in 2 patients. Three (5.9%) of the 51 patients who underwent elective operation and 12 (63.2%) of the 19 patients with emergency operation died in the hospital, giving an overall hospital mortality of 21.4% (15/70). Hospital mortality was reduced from 33.3% (8/24) in the initial 24 patients to 15.2% (7/46) in the recent 46 patients with the volume reduction test. Mean ejection fraction increased from 22.2 +/- 6.7% to 29.6 +/- 6.0%. Diastolic dimension decreased from 81.1 +/- 9.5 to 69.8 +/- 19.2 mm. End-diastolic and systolic volume indices decreased from 199.0 +/- 47.9 to 124.1 +/- 34.9 ml/m2 and from 154.0 +/- 41.2 to 89.3 +/- 31.7 ml/m2, respectively, at one postoperative month in the 55 hospital survivors. The mean pulmonary capillary wedge pressure decreased from 25.6 +/- 7.8 to 13.6 +/- 4.5 mmHg. Serum brain natriuretic peptide decreased from 999 +/- 647 preoperatively to 547 +/- 362 pg/ml one month after the operation. Thirteen patients (18.6%) died in the late period mainly due to heart failure. Among the 42 survivors, 37 patients returned to NYHA functional class I-II, and cardiac events were rare after one postoperative year. Actuarial survivals at 3 years in elective and emergency operations were 71.9% and 33.3%, respectively. CONCLUSIONS: Left ventriculoplasty is acceptably safe for elective operation and clinical improvement can be obtained by proper surgical procedures and careful postoperative medical treatment.

[Combined valve and coronary artery bypass surgery].

Between January 1997 and November 1999, 31 patients underwent combined valve and coronary artery bypass grafting (CABG) surgery in our institute. There was no hospital death and clinical symptom improved in all patients after surgery. Mean follow-up was 16.3 months. The actuarial survival rate was 96.8% at 1 year. There were 4 late deaths (CHF in 3, arrhythmia in 1). Tepid blood cardioplegia was effective for those operations. Operation of CABG combined with valve surgery or LV surgery could be safely performed. We therefore considered that complete correction of the lesion with not only CABG but also combined valve surgery or LV surgery was the procedures of choice in patients with complicated lesions.

[Aortic valve/root replacement using a stentless bioprosthesis (Medtronic Freestyle valve)].

BACKGROUND: The purpose of the study is to assess the clinical and hemodynamic performance of aortic valve replacement (AVR) with the Freestyle bioprosthesis. METHODS: Twenty-one patients received AVR with a Freestyle aortic root bio-prosthesis between May 1998 and October 1999. Eighteen patients underwent AVR with subcoronary method and three patients with aortic root (full root) method. Patients were evaluated postoperatively at discharge by clinical examination and color Doppler echocardiography. RESULTS: There was one death due to multi-organ failure. No patients experienced valve deterioration, paravalvular leak, unacceptable hemodynamic performance, nor thromboembolic event. Excellent function is demonstrated by very low gradient (mean gradient 7.2 +/- 4.7 mmHg) through aortic valve and no significant aortic regurgitation (none: 11, trivial/mild: 10). All patients had been in New York Heart Association Functional Class III and IV preoperatively, and after surgery, 17 patients were in Class I, and 3 were in Class II. CONCLUSION: The Freestyle bioprosthesis has good clinical and hemodynamic performance without Coumadin. Further follow-up is required to evaluated valve durability.

Minimally invasive coronary artery revascularization: off-pump bypass grafting and the hybrid procedure.

was safe, effective, and suitable especially in patients with high risks for coronary artery bypass grafting.

Cerebral acetylcholine levels and long-term spatial cognitive impairment following middle cerebral artery occlusion in rats.

The behavioral and neurochemical changes in the chronic phase of permanent occlusion of the right middle cerebral artery (MCA) in rats were investigated. Nineteen MCA-occluded rats failed to solve the 8-arm radial maze task (cognitively impaired rats), while 11 MCA-occluded rats could complete it (cognitively unimpaired rats). When a delay of 60 min was imposed in the task, however, 5 cognitively unimpaired rats failed to complete the task. The rats that underwent behavioral testing were studied for any changes in ACh levels in various brain regions using HPLC with electrochemical detection. The ACh levels in the infarcted areas decreased considerably in all MCA-occluded rats, but no region of the infarcted areas correlated with the spatial cognitive deficit. The ACh levels tended to decrease in the frontal cortex of the cognitively impaired rats and greatly increased in both ipsilateral and contralateral parietal cortex of the cognitively unimpaired rats. A significant correlation was observed between the ACh levels and spatial cognitive deficit in the contralateral frontal cortex, and ipsilateral and contralateral parietal cortex. These results suggest that the cholinergic function of the frontal and parietal cortices might play a role in acquiring spatial cognition in MCA-occluded rats.

[Determinants of long-term outcome for operative survivors of acute aortic dissection].

Predictability of aorta-related complications and survival was examined in 79 operative survivors of acute aortic dissection. Follow-up was 94.9% complete and totaled 458 patient-years. Actuarial survival was 93 +/- 3% (+/- S.E.) (n = 43) at 5 years, and 74 +/- 8% (n = 13) at 10 years. Survival was significantly lower in patients having neurological complication. Freedom from aorta-related complications was 82 +/- 5% (n = 37) at 5 years, and 67 +/- 8% (n = 11) at 10 years. Multivariate Cox regression analysis identified residual entry and leak on anastomotic site as independent predictors of aorta-related complications. We conclude that in the treatment of acute aortic dissections, reducing the incidence of residual entry and leak on anastomotic site improves long-term outcome.

Enhanced protein synthesis in the ipsilateral substantia nigra following middle cerebral artery occlusion in the rat.

Following focal cerebral ischemia, neuronal cell death is detected in remote areas of the brain, including the ipsilateral thalamus and substantia nigra (SN), as well as in the ischemic core. We have investigated protein synthesis in the remote areas of rats exposed to focal ischemia using autoradiography. The proximal portion of the left middle cerebral artery (MCA) was permanently occluded, and at various periods (6 h, 2, 4 and 7 days and 2 and 4 weeks following ischemia) animals received a single dose of L-[2,3-3H]valine (6.7 mCi/kg). Brain sections containing the thalamus and SN were processed for autoradiography. In the ipsilateral cerebral cortex and striatum, marked impairment of protein synthesis was observed and was never completely recovered during the experiment. No changes in protein synthesis in the ipsilateral thalamus were detected during the experiment. However, a change in protein synthesis was demonstrated in the ipsilateral SN. At 2 days after MCA occlusion, incorporation of [3H]valine into the whole zona reticulata of the ipsilateral SN was slightly enhanced and the increase became evident at 4 days after ischemia. Increased incorporation of [3H]valine began to be localized in the lateral portion of the zona reticulata after 7 days and continued up to 4 weeks following ischemia. Enhanced protein synthesis during the early stage (2 and 4 days after ischemia) may be due to the activated function of the neurons in the zona reticulata and that during the late stage (7 days and 2 and 4 weeks) after ischemia to astroglial proliferation

[Changes in acetylcholine level and its related enzyme activities in rat brain following focal ischemia].

At 1 to 4 weeks following middle cerebral artery occlusion in Wistar rats, levels of acetylcholine, which is neuro-transmitter, were measured. In the hippocampus, which plays an important role in memory, levels of acetylcholine in the ischemic group could not be detected the significant difference compared with those in the sham-operated group. But in the anterior cortex and the caudate-putamen, levels of acetylcholine in the ischemic group were recognized the significant difference compared with those in the sham operated group. Moreover activities of choline acetyltransferase, which is the synthetic enzyme of acetylcholine, and activities of acetylcholinesterase, which is the degradative enzyme of acetylcholine, were measured in the anterior cortex which was recognized decrease of the acetylcholine level and the hippocampus which could not be detected the difference of the acetylcholine level. Activities of both enzymes in the hippocampus could not be recognized the significant difference between the ischemic group and the normal group. But in the anterior cortex activities of both enzymes in the ischemic group were significantly decreased compared with those in the normal group. These results suggest that these decreases in the ischemic group were due to damage of injection of cholinergic neuron from Meynert nucleus (basal nucleus of Meynert) to anterior cortex.

Complement activation by diesel exhaust particles (DEP).

The effect of diesel exhaust particles (DEP) on the hemolytic activity of human serum complement was investigated. Previous treatment of human serum with DEP extracts at 37 degrees C decreased the hemolytic activity of human serum complement dose dependently, to 20% of its original value. This decrease in complement activity by DEP extracts was observed with previous incubation at 37 degrees C but not at 4 degrees C. A decrease in complement activity was observed after previous incubation at 37 degrees C in the presence of EGTA/Mg but not in the presence of EDTA, indicating that the alternative pathway of the complement system had been activated by DEP extracts. Activation of the complement system by DEP extracts was further demonstrated by observation of the cleavage of the third component of the complement (C3) in serum to C3b with immunoelectrophoresis using goat anti-human C3 antiserum. This cleavage of C3 was similarly observed in the presence of EGTA/Mg, indicating the activation of the alternative pathway of the complement system by DEP extracts. These results indicate that DEP can activate the alternative pathway of the complement system, resulting in a decrease in the hemolytic activity of complement and in the production of biologically active degradation products of complement proteins such as C3a. The biological significance of the activation of the complement by DEP in the alveolus is discussed in relation to the influx of neutrophiles.

[The case of left atrial myxoma originating from posterior leaflet of mitral valve].

We experienced the case of left atrial myxoma originating from posterior leaflet mitral valve. Mitral valve repair was performed in that case, because of the presence mitral leaflet defect due to removal of the myxoma. Transesophageal echocardiography was a useful tool for perioperative evaluation of the mitral valve and precise localization of the origin of the myxoma.

Effect of L-arginine and NG-nitro-L-arginine on delayed neuronal death in the gerbil hippocampus.

To assess the role of nitric oxide (NO) in cerebral ischemia, we investigated the effect of L-arginine, a substrate of NO synthase (NOS), and NG-nitro-L-arginine (L-NNA), a NOS inhibitor, on neuronal death in the CA1 hippocampal region. Seventy-two Mongolian gerbils were used in the study. Both carotid arteries were occluded for 4 min to induce forebrain ischemia. Temporal muscle temperature was strictly maintained at 37.5 +/- 0.3 degrees C during the ischemia. L-arginine (10 and 100 mg kg-1) or L-NNA (1, 10 and 100 mg kg-1) was administered intraperitoneally 4 times: 30 min before, 3 h, 6 h and 24 h after induction of ischemia. Four days after ischemic insult, the animals were perfusion-fixed, and the neuronal densities in the medial, middle and lateral CA1 subfield were estimated. Average neuronal cell density of the control group was 2-3 mm in each subfield. L-arginine at doses of 10 and 100 mg kg-1 did not prevent neuronal death. L-NNA at doses of 1 and 10 mg kg-1 did not protect neuronal cells from ischemia either. However, in ischemia gerbils treated with 100 mg kg-1 L-NNA, the average neuronal cell density in the lateral CA1 subfield was 54.4 +/- 19.1, L-NNA (100 mg kg-1) significantly (p < 0.05) reduced the occurrence of neuronal death in the lateral CA1 subfield. The present results suggest that NO plays an important role in the development of neuronal injury after global ischemia.