[Integrated coronary artery bypass strategy prevents urgent pump conversion during off-pump coronary artery bypass grafting].

Urgent pump conversion during off-pump coronary artery bypass (OPCAB) results in high morbidity and mortality. We retrospectively evaluated if the peri-operative integrated strategy prevents this lethal event in our 400 consecutive OPCAB operations. The patients with preoperative cardiogenic shock and/or ventricular arrhythmias underwent on-pump coronary artery bypass grafting (CABG). All other patients (99% of total CABG) were scheduled to undergo OPCAB (n=400). Prophylactic intraaortic balloon pumping (IABP) was applied to the patients with critical (>95%) left main trunk stenosis or low (<0.35) left ventricular ejection fraction. All the patients received the deep pericardial suture, apex-traction device, suction-type stabilizer, test-clamp of target coronary arteries by micro bulldog clamp, and intra-coronary shunts. Intra-operative IABP was applied in the case of sustained ST-segment change and/or elevated pulmonary artery pressure. Pump conversion was indicated for the patients with ventricular fibrillation and/or cardiogenic shock. Two patients (0.5%) had pump conversion due to ventricular arrhythmia and sustained hypotension, respectively. These pump conversion did not result in hospital mortality. Three hospital deaths (0.7%) occurred due to non-cardiac causes. The integrated strategy using prophylactic or intra-operative IABP in OPCAB produce a low pump conversion rate even during an early period of surgeon's learning curve.

Inhibition of the spindle assembly checkpoint kinase Mps-1 as a novel therapeutic strategy in malignant mesothelioma.

Malignant mesothelioma (MM) is an aggressive malignancy, highly resistant to current medical and surgical therapies, whose tumor cells characteristically show a high level of aneuploidy and genomic instability. We tested our hypothesis that targeting chromosomal instability in MM would improve response to therapy. Thr/Tyr kinase (TTK)/monopolar spindle 1 kinase (Mps-1) is a kinase of the spindle assembly checkpoint that controls cell division and cell fate. CFI-402257 is a novel, selective inhibitor of Mps-1 with antineoplastic activity. We found that CFI-402257 suppresses MM growth. We found that Mps-1 is overexpressed in MM and that its expression correlates with poor patients' outcome. In vitro, CFI-402257-mediated inhibition of Mps-1 resulted in abrogation of the mitotic checkpoint, premature progression through mitosis, marked aneuploidy and mitotic catastrophe. In vivo, CFI-402257 reduced MM growth in an orthotopic, syngeneic model, when used as a single agent, and more so when used in combination with cisplatin+pemetrexed, the current standard of care. Our preclinical findings indicate that CFI-402257 is a promising novel therapeutic agent to improve the efficacy of the current chemotherapeutic regimens for MM patients.

DNA methylation and expression of the rat pepsinogen gene in embryonic, adult, and neoplastic tissues.

The relationship between methylation and expression of rat pepsinogen 1 (Pg1) genes was investigated in various tissues. On Northern blotting with a Pg1 complementary DNA probe, Pg1 mRNA was detected only in the glandular stomach of normal rats. Methylation analysis with Msp1/HpaII and Hha1 revealed tissue specific methylation patterns of Pg1 genes with less methylated in the stomach than in other normal tissues not expressing the genes. During stomach development, there was a progressive increase in the Pg1 mRNA level that almost coincided with change in the mucosal pepsinogen level and progressive demethylation after the onset of transcription. Thus, there was an inverse correlation between methylation and expression of Pg1 genes, suggesting a role of DNA methylation in Pg1 gene regulation during normal differentiation, although not its primary role in gene activation. There was no detectable Pg1 mRNA in either primary or transplanted stomach cancers induced by N-methyl-N'-nitro-N-nitrosoguanidine. The methylation patterns of Pg1 genes were different from those of normal tissues that expressed the gene and of those that did not and no simple correlation was observed between methylation and expression of Pg1 genes. This result is consistent with a previous finding that DNA methylation is deranged in tumor cells.

Minimal asbestos exposure in germline BAP1 heterozygous mice is associated with deregulated inflammatory response and increased risk of mesothelioma.

Germline BAP1 mutations predispose to several cancers, in particular malignant mesothelioma. Mesothelioma is an aggressive malignancy generally associated with professional exposure to asbestos. However, to date, we found that none of the mesothelioma patients carrying germline BAP1 mutations were professionally exposed to asbestos. We hypothesized that germline BAP1 mutations might influence the asbestos-induced inflammatory response that is linked to asbestos carcinogenesis, thereby increasing the risk of developing mesothelioma after minimal exposure. Using a BAP1(+/-) mouse model, we found that, compared with their wild-type littermates, BAP1(+/-) mice exposed to low-dose asbestos fibers showed significant alterations of the peritoneal inflammatory response, including significantly higher levels of pro-tumorigenic alternatively polarized M2 macrophages, and lower levels of several chemokines and cytokines. Consistent with these data, BAP1(+/-) mice had a significantly higher incidence of mesothelioma after exposure to very low doses of asbestos, doses that rarely induced mesothelioma in wild-type mice. Our findings suggest that minimal exposure to carcinogenic fibers may significantly increase the risk of malignant mesothelioma in genetically predisposed individuals carrying germline BAP1 mutations, possibly via alterations of the inflammatory response.

Aspirin delays mesothelioma growth by inhibiting HMGB1-mediated tumor progression.

High-mobility group box 1 (HMGB1) is an inflammatory molecule that has a critical role in the initiation and progression of malignant mesothelioma (MM). Aspirin (acetylsalicylic acid, ASA) is the most widely used nonsteroidal anti-inflammatory drug that reduces the incidence, metastatic potential and mortality of many inflammation-induced cancers. We hypothesized that ASA may exert anticancer properties in MM by abrogating the carcinogenic effects of HMGB1. Using HMGB1-secreting and -non-secreting human MM cell lines, we determined whether aspirin inhibited the hallmarks of HMGB1-induced MM cell growth in vitro and in vivo. Our data demonstrated that ASA and its metabolite, salicylic acid (SA), inhibit motility, migration, invasion and anchorage-independent colony formation of MM cells via a novel HMGB1-mediated mechanism. ASA/SA, at serum concentrations comparable to those achieved in humans taking therapeutic doses of aspirin, and BoxA, a specific inhibitor of HMGB1, markedly reduced MM growth in xenograft mice and significantly improved survival of treated animals. The effects of ASA and BoxA were cyclooxygenase-2 independent and were not additive, consistent with both acting via inhibition of HMGB1 activity. Our findings provide a rationale for the well documented, yet poorly understood antitumorigenic activity of aspirin, which we show proceeds via HMGB1 inhibition. Moreover, the use of BoxA appears to allow a more efficient HMGB1 targeting while eluding the known gastrointestinal side effects of ASA. Our findings are directly relevant to MM. Given the emerging importance of HMGB1 and its tumor-promoting functions in many cancer types, and of aspirin in cancer prevention and therapy, our investigation is poised to provide broadly applicable information.

Cerebral metabolic recovery from deep hypothermic circulatory arrest after treatment with arginine and nitro-arginine methyl ester.

BACKGROUND: Recent studies suggest that nitric oxide is important in the pathogenesis of ischemic brain injury and also has a role in controlling cerebrovascular tone. This study examines the net effects of nitric oxide on cerebral metabolic recovery after deep hypothermic circulatory arrest. METHODS: Two-week-old piglets were supported by cardiopulmonary bypass and cooled to 15 degrees C followed by 1 hour of deep hypothermic circulatory arrest, 45 minutes of reperfusion and rewarming, and then 3 hours of normothermic perfusion. Groups of 10 piglets received one of four treatments before bypass; L-nitro-arginine methyl ester, inhibitor of nitric oxide synthesis, 10 mg/kg intravenously; L-arginine, to enhance nitric oxide synthesis, 30 mg/kg intravenously before bypass and then 10 mg/kg per minute during the first hour of reperfusion; a combination of L-nitro-arginine methyl ester plus L-arginine at these same doses; and no pretreatment (controls). Cerebral high-energy phosphates and pH were measured by magnetic resonance spectroscopy in half the animals. Cerebral blood flow, metabolic rates for oxygen and glucose, and the oxidation/reduction state of cytochrome aa3 and oxygenated and deoxygenated hemoglobin measured by near-infrared spectroscopy were assessed in the other half of the piglets. RESULTS: L-nitro-arginine methyl ester significantly increased cerebral vascular resistance and markedly reduced recovery of high-energy phosphates, pH, and oxidation state of cytochrome aa3, L-arginine increased cerebral blood flow, cerebral glucose and oxygen consumption, and recovery of cytochrome aa3 oxidation and high-energy phosphates. L-Arginine did not reverse completely the effects of L-nitro-arginine methyl ester on cerebral metabolic recovery. CONCLUSION: In a piglet model of deep hypothermic circulatory arrest, L-nitro-arginine methyl ester has a deleterious effect and L-arginine has a beneficial effect on cerebral metabolic recovery. The deleterious metabolic effects of L-nitro-arginine methyl ester are only partially reversed by L-arginine. This fact suggests that there may be mechanisms in addition to inhibition of nitric oxide synthesis contributing to the neurotoxicity of L-nitro-arginine methyl ester in this model.

The leukemic oncogene tal-2 is expressed in the developing mouse brain.

tal-1 (T-cell acute leukemia-1; also known as SCL) and tal-2 genes belong to a family of basic helix-loop-helix transcription factors and were originally isolated from the breakpoints of chromosomal translocations in human T-cell leukemia cell lines. tal-1 is expressed not only in hematopoietic cells but also in several endothelial structures and the central nervous system during development. On the other hand, the detailed function and the sites of expression of tal-2 have remained obscure. We cloned the tal-2 cDNA from a mouse embryonic cDNA library and examined its expression pattern in the mouse, comparing with that of tal-1. In situ analyses revealed that tal-2 transcripts are detected at embryonic day 12.5 in the following regions; 1) the diencephalon-the zona limitans intrathalamica and the pretectum, 2) the mesencephalon-the tectum, and the anterior and posterior tegmentum, 3) the metencephalon-the isthmus and the anterior pons. In the diencephalon and the mesencephalon, the expression sites of tal-2 gene were similar to those of tal-1, and its expression was stronger than that of tal-1. In the metencephalon, tal-2 expression was observed in the anterior pons, whereas tal-1 transcripts were detected in the entire pons, and showed stronger expression than tal-2. The tal-2 messages were barely detectable in the brain at birth. These results suggest that tal-1 and tal-2 are involved in the development of specific areas of the central nervous system.

The primary structure of the major pepsinogen from the gastric mucosa of tuna stomach.

The complete primary structure of the major component of tuna pepsinogens was determined by conventional protein chemistry methods. It was composed of a prosegment of 37 residues and a pepsin moiety of 323 residues, having a relative molecular mass of 39,364. The essential aspartyl residues in the active site and the three disulfide bonds common to other pepsinogens were conserved; however, several unique substitutions and/or deletions characteristic of tuna pepsinogen were found at various positions, especially in the prosegment and subsite regions, as compared with the sequences of other pepsinogens, which may affect the rate of activation of the zymogen, and/or the catalytic function and substrate specificity of the enzyme. Tuna pepsinogen is the least acidic among pepsinogens. The sequence identity between tuna pepsinogen and other pepsinogens ranged from 45 to 52%. A phylogenetic tree based on the primary structures suggested that tuna pepsinogen diverged from the pepsinogen A and prochymosin groups in an early period of pepsinogen evolution.

A comparative study on the NH2-terminal amino acid sequences and some other properties of six isozymic forms of human pepsinogens and pepsins.

Six pepsinogen isozymogens, including five forms of pepsinogen A (PGA) and an apparently single form of pepsinogen C (PGC), were isolated simultaneously from the purified total pepsinogen fraction of human gastric mucosa by fast protein liquid chromatography on a Mono Q column, and their NH2-terminal amino acid sequences and some other properties were compared. Upon activation at pH 2.0, all the isozymogens were converted to the corresponding pepsins in a stepwise manner through intermediate forms. The activation rates and the cleavage sites in the activation peptide segment to generate intermediate forms were significantly different among the isozymogens. The NH2-terminal 85-residue amino acid sequences of these isozymogens were determined, including the sequences of the activation peptide segments and the NH2-terminal regions of the corresponding pepsins. Differences in amino acid sequence were found at positions 43 and 77 among the pepsinogen A isozymogens; the residue at position 43 was Lys in PGA-5, PGA-4, and PGA-3a, and Glu in PGA-3 and PGA-2, and the residue at position 77 was Leu in PGA-5 and PGA-4 and Val in PGA-3 and PGA-2. Phosphate was not found in any of the isozymogens. The corresponding pepsins also showed significant variations in properties such as specific activities toward synthetic and protein substrates, pH dependence of activity, susceptibility to various inhibitors, and thermal and alkaline stabilities.

Difference of activation processes and structure of activation peptides in human pepsinogens A and progastricsin.

The activation processes of two human pepsinogens A (pepsinogens 3 and 5) and progastricsin were compared with special attention to pepsinogens 3 and 5. Each zymogen was converted to pepsin in a stepwise manner through intermediate forms. In pepsinogens A, the major cleavage site was the Leu23-Lys24 bond and this cleavage was suggested to occur intramolecularly. When each of the pepsins A was added to the corresponding pepsinogen A exogenously, the latter was rapidly converted to pepsin, releasing the 47-residue intact activation segment. In this case, the Leu47-Val48 bond connecting the activation segment with the pepsin moiety was cleaved by an intermolecular reaction. On the other hand, when the pepsinogen A-pepstatin complex was attacked by each corresponding pepsin A added exogenously, significant cleavage by an intermolecular reaction occurred at the Asp25-Phe26 bond, generating the Phe26-intermediate form. These shifts of the cleavage sites in pepsinogens A depending on the activation conditions are likely to correlate with the conformation of the activation segment. These results can be explained consistently in terms of a proposed molecular model of activation.

Probing stability and dynamics of proteins by protease digestion. II: Identification of the initial chymotryptic cleavage sites of homologous cytochromes c.

Three homologous cytochromes c from horse, rabbit and tuna were subjected to chymotryptic digestion and their initial cleavage sites were identified. The sites in oxidized cytochromes c are the COOH-terminal sides of Tyr-48, Phe-46 and Tyr-46 for horse, rabbit and tuna cytochromes c, respectively. The results show that the chymotrypsin attacks a single site in each protein; the sites are located at the almost identical position on the polypeptide chain. Through the time-course studies of digestion, it was found that the three cytochromes c have different chymotrypsin-susceptibility at the initial cleavage site in the order of horse less than rabbit less than tuna. Studies on chymotryptic digestion of tuna cytochrome c in the reduced form revealed that the haem-reduction does not alter the initial cleavage site but increases the resistance to the proteolysis at the site. The uniqueness of the initial cleavage site in each cytochrome c species suggests that the protease susceptibility reflects some overall properties of the protein. At the same time, it was clarified that the initial cleavage site is also affected by a neighboring region by the fact that another potential cleavage site is located near the site in question. In order to elucidate the initial cleavage site, several physical properties of tuna cytochrome c molecule deduced from the X-ray 3D structure, accessible surface area, temperature factor, effective hydrophobicity and electrostatic potential, were compared with the experimental results and it was concluded that these properties given by a residue have no direct relationship with the chymotrypsin susceptibility.

A steroid-binding protein mediates estrogen-dependent inhibition of growth of MCF-7 breast cancer cells.

We have described two different proteins that mediate MCF-7 cell growth inhibition (1). One is estrogen-dependent and is a heterodimer consisting of a heavy (54 kDa) and light (29 kDa) chain. This protein is distinct from cortisol-binding globulin (CBG), sex hormone-binding protein (SBP or SHBG) and albumin (HSA). It may be fetal steroid binding protein (FSBP). The other is not estrogen dependent, but may be related to the estrogen-dependent protein. Since these are large proteins they must be acting at the level of the plasma membrane, not traditional intracellular estrogen receptors, and inhibit MCF-7 cell growth.

Growth inhibition of MCF-7 cells by estrogen is dependent upon a serum factor.

MCF-7 cell growth is normally dependent upon estrogen, but if cells are maintained in serum-free medium estrogen inhibits cell growth with an IC50 of about 1 nM. Cells adapted to serum-free medium have approximately the same levels of estrogen receptor mRNA as control cells that are stimulated by estrogen. We have identified two serum components, one estrogen dependent and one not, that appear to be responsible for the inhibition of growth by estrogen. Our observations are consistent with the presence of a plasma membrane receptor which negatively regulates MCF-7 cell growth, and that can be inhibited by a serum protein that binds estrogen.

Inhibitory effect of cimetidine on graft survival of allotransplanted rats submitted to an active-passive enhancement protocol.

The objective of the present study was to determine whether cimetidine, a type-2 histamine receptor antagonist, inhibits the immunological enhancement of allografted rats achieved by treatment with donor antigen plus anti-donor antibody. Groups of rats submitted to this active-passive enhancement protocol and treated ip with 30 (APEC 30; Group I; N = 4) or 60 (APEC 60; Group II; N = 8) mg/day cimetidine for 14 days had a significantly shorter graft survival (20.2 +/- 5.1 and 11.1 +/- 2.6 days, respectively) than the control group (animals submitted to the enhancement protocol and killed on day 72 after transplant when the graft was beating normally; APE; Group III; N = 6; P less than 0.05). On the other hand, these animals had a significantly longer graft survival than rats allotransplanted but not treated for enhancement (ALLO; Group V; N = 5; 8.2 +/- 0.8 days). The surgical control, consisting of isotransplanted animals, had a long-term survival (ISO; Group VI; N = 6; rats killed 120 days after transplant with the graft beating normally). Animals treated with cimetidine, but not submitted to the enhancement protocol (AC 60; Group IV, N = 4) had a significantly shorter graft survival (6.25 +/- 0.5) than the allotransplanted animals (Group V). These results indicate inhibition of the suppressor mechanisms which participate in this type of immunological enhancement.

Immunomodulatory effect of cimetidine on the proliferative responses of splenocytes from T. cruzi-infected rats.

The immunomodulatory effect of cimetidine (CIM), a histamine type-2 receptor antagonist, was evaluated in respect to the blastogenic response to Con A of Wistar Furth (WF) rats infected by the Y strain of Trypanosoma cruzi (T. cruzi). Enhancement of blastogenesis of normal splenocytes was observed at a concentration of 10(3) M. However, the splenocytes from infected animals responded to concentrations of CIM ranging from 10(-8) to 10(-3) M. The mitogenic response to Con A of cells from infected animals was restored in the presence of CIM. The results show that CIM modulates the "in vitro" proliferative response of cells from T. cruzi-infected rats and suggest an immunoregulatory role of histamine and/or of cells that express H2 receptors in this infection.

HIV heterosexual transmission to stable sexual partners of HIV-infected Brazilian hemophiliacs.

Nineteen Brazilian HIV-infected hemophiliacs and their stable heterosexual sexual partners were studied with the aim of assessing the rate of HIV transmission in this at risk group. The mean length of relationship between couples was 7.4 years. The hemophiliac men were Class II (n = 6), III (n = 11) and IVa (n = 2) of the CDC classification. They had decreased CD4+ and elevated CD8+ cell numbers; five had p24 antigenemia. We found 3 HIV-infected women (15.8 percent) by routine and confirmatory tests, a prevalence similar to that seen in other countries. They were asymptomatic and had no detectable p24 antigenemia. The 3 seropositive women's partners were Class II and III-CDC, and had normal CD4+ and CD8+ values and no p24 antigenemia. All seronegative women also had normal CD4+ and CD8+ numbers, except for elevated CD8+ cells in three of them, but immune abnormalities had already been seen in some seronegative partners at high risk for HIV infection. Our results reinforce previous suggestions that heterosexual transmission to stable female partners occurs preferentially early after initiation of sexual exposure, and possibly when the transmitter had high levels of viremia and regular sexual activity.

PIP: In Brazil, hemophiliacs who received coagulation factor concentrates during 1980-85 have rates of HIV exceeding 50% and rates of heterosexual transmission to their partners in the range of 10-20%. This study investigated the clinical course of HIV infection in 19 male patients from a hematology center in Sao Paulo, Brazil, with hemophilia A or B and their stable, asymptomatic female sexual partners. The mean duration of the relationship was 7.4 years. Compared with 15 normal adult subjects used as controls, CD8+ cell counts of hemophiliacs were significantly higher while CD4+ cell values were significantly reduced. Three sexual partners (15.8%) were HIV-positive, implying a transmission rate of 2.1 per 100 person-years. All female partners were in Centers for Disease Control Class II. Their male partners were in Classes II and III and had normal CD4+ and CD8+ levels. Neither males nor females had p24 antigenemia. Fragments of HIV particles were present in several HIV-negative female partners. These findings suggests early HIV transmission, when the transmitter has high levels of viremia, to stable female partners of hemophiliacs.

[Two cases of atrial septal defect with absence of right superior vena cava and persistent left superior vena cava].

Two women (16-year-old, 54-year-old) of atrial septal defect with absence of right superior vena cava and persistent left superior vena cava were successfully operated on. Electrocardiographic findings show coronary sinus rhythm and atrial fibrillation. When we closed atrial septal defect cannulated to persistent left superior vena cava via the coronary sinus directly, sick sinus syndrome was not appeared postoperatively.

[Left main coronary artery stenosis following aortic valve replacement using a solid coronary perfusion catheter: report of two cases].

We experienced two cases of iatrogenic left main coronary artery stenosis (IOCS) following double (aortic and mitral) valve replacement (DVR). The solid coronary perfusion catheter may attribute IOCS, with grave consequence. There have been no IOCS since the time we exchanged a solid catheter for a soft one. One case, she was successfully treated percutaneous transluminal coronary angioplasty (PTCA), because she developed angina pectoris about 5 years after PTCA. But she developed angina pectoris again and angiographically left main coronary was severe stenotic. So she was undergone aorto coronary bypass grafting (CABG) to the left anterior descending. The other case, he developed angina pectoris about 3 months after DVR. He was treated with PTCA. Angiographically left mine coronary artery stenosis reduced 50% from 90%. Generally the treatment of IOCS is CABG, but we performed PTCA for 2 patients. Because we thought it was very hazardous for us to perform them open heart surgery. When it is very hazardous to perform patients open heart surgery, they need to be performed PTCA.

[A case of ruptured aneurysm of Valsalva sinus into right atrium with peculiar findings on aortography].

A 24-year-old man with ruptured aneurysm of sinus of Valsalva into the right atrium originating from the noncoronary sinus is presented. On aortography through the ascending aorta the right atrium in systolic phase and the right ventricle in diastolic phase were opacified. We considered ruptured aneurysm like a streamer (wind sock) entered into the right ventricle in diastolic phase and into the right atrium in systolic phase. Post-aneurysmectomy course was uneventful, and radiographic examination revealed complete repair of the aneurysm.

[A case report of perforated aneurysm of mitral valve with aortic regurgitation].

The patient was a 71-year-old male who complained of palpitation and tachycardia. The echocardiogram showed a bulging of the anterior mitral valve leaflet toward the left atrium that persisted throughout cardiac cycle. The cine angiogram showed deformity of the anterior mitral valve leaflet with severe mitral regurgitation and mild aortic regurgitation. At operation, a perforated aneurysm was recognized at the anterior mitral valve leaflet without thrombus and vegetation. The size of aneurysm was 40 x 25 x 25 mm. The patient underwent MVR + AVR, and the postoperative course was uneventful. Pathological examination of the anterior mitral valve leaflet revealed scar-like fibrosis and old inflammatory change. It was judged a true aneurysm of mitral valve, because the structure of endocardium was kept.