Impact of bridging with perioperative low-molecular-weight heparin on cardiac and bleeding outcomes of stented patients undergoing non-cardiac surgery.

When patients with coronary stents undergo non-cardiac surgery, bridging therapy with low-molecular-weight heparin (LMWH) is not infrequent in clinical practice. However, the efficacy and safety of this approach is poorly understood. This was a retrospective analysis of patients with coronary stent(s) on any antiplatelet therapy undergoing non-cardiac surgery between March 2003 and February 2012. The primary efficacy endpoint was the 30-day incidence of major adverse cardiac or cerebrovascular events (MACCE), defined as the composite of cardiac death, myocardial infarction, acute coronary syndrome leading to hospitalisation, or stroke. The primary safety endpoint was the 30-day composite of Bleeding Academic Research Consortium (BARC) bleedings ≥ 2. Among 515 patients qualifying for the analysis, LMWH bridging was used in 251 (49 %). At 30 days, MACCE occurred more frequently in patients who received LMWH (7.2 % vs 1.1 %, p=0.001), driven by a higher rate of myocardial infarction (4.8 % vs 0 %, p< 0.001). This finding was consistent across several instances of statistical adjustment and after the propensity matching of 179 pairs. Patients bridged with LMWH also experienced a significantly higher risk of BARC bleedings ≥ 2 (21.9 % vs 11.7 %, p=0.002) compared to those who were not, which remained significant across different methods of statistical adjustment and propensity matching. In conclusion, LMWH bridging in patients with coronary stents undergoing surgery is a common and possibly harmful practice, resulting in worse ischaemic outcomes at 30 days, and a significant risk of bleeding.

Mitral valve repair and transesophageal echocardiographic findings in a high-risk subgroup of patients with active, acute infective endocarditis.

BACKGROUND AND AIM OF THE STUDY: Limited data are available regarding the efficacy of mitral valve repair in patients affected by active, acute infective endocarditis. In addition, the predictivity of transesophageal echocardiography (TEE) for guiding the surgical decision-making process in these patients has not yet been reported. The study aim was to evaluate the long-term results of mitral valve repair and role of TEE in active, acute infective endocarditis. METHODS: The study population consisted of patients affected by infective endocarditis of the mitral valve who underwent surgery. TEE was performed intraoperatively to guide the best surgical approach. All patients were followed up (mean 73+/-8 months) after surgery. RESULTS: Twenty-eight patients underwent surgery for infective endocarditis; of these, 13 had mitral valve repair for active, acute infective endocarditis and formed the basis of the study. Sensitivity, specificity, positive predictive value, negative predictive value of TEE in detecting the mechanism of mitral regurgitation were 87%, 100%, 100% and 92%, respectively. The predictivity test of TEE in guiding surgical strategy was 94%. All patients were alive at the time of follow up; 10 (77%) were in NYHA class I and three in class II (23%). Mitral regurgitation was severe in one patient (8%), moderate in three (23%), mild in four (31%), and absent in five (38%). No relapses of active infective endocarditis were observed during the follow up period. CONCLUSION: Mitral valve repair appears to be an effective treatment for active, acute infective endocarditis with mitral regurgitation and should be considered as a therapeutic strategy when surgery is contemplated. TEE has a fundamental role in the surgical decision-making process in these patients.

Repair of giant hernias using more prosthesis.

Giant incisional hernias with total loss of substance are an ominous pathological condition characterized by massive depletion of muscular and fascial tissue, by complete loss of the anatomical and physiological function of the abdominal wall and by severe respiratory and visceral involvement. Over a 10-year period we operated 270 patients with voluminous incisional hernias, 12 of which had a total loss of substance. There was no intraoperative mortality. One patient died of myocardial infarction on the fifth and one died of intestinal occlusion and peritonitis the 11th postoperative day. Early postoperative complications occurred in only one patient who had skin necrosis with an infection at the polypropylene mesh. This was successfully treated with systemic antibiotic therapy and topical medication of the wound. There was also one minor recurrence over the pubis 1 year after the operation that required a new operation to replace the mesh. No respiratory complications occurred and all patients were normally active. The good results reported in our series encourage us to continue in this direction even though these patients are at high risk.

TGF-beta autocrine loop regulates cell growth and myogenic differentiation in human rhabdomyosarcoma cells.

Transforming growth factor beta (TGF) is a well-known inhibitor of myogenic differentiation as well as an autocrine product of rhabdomyosarcoma cells. We studied the role of the TGF-beta autocrine loop in regulating growth and myogenic differentiation in the human rhabdomyosarcoma cell line, RD. We previously reported that the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) induces growth arrest and myogenic differentiation in these cells, which constitutively express muscle regulatory factors. We show that TPA inhibits the activation of secreted latent TGF-beta, thus decreasing the concentration of active TGF-beta to which the cells are exposed. This event is mediated by the TPA-induced alteration of the uPA/PAI serine-protease system. Complete removal of TGF-beta, mediated by the ectopic expression of a soluble type II TGF-beta receptor dominant negative cDNA, induces growth arrest, but does not trigger differentiation. In contrast, a reduction in the TGF-beta concentration, to a range of 0.14-0.20 x 10(-2) ng/ml (which is similar to that measured in TPA-treated cells), mimics TPA-induced differentiation. Taken together, these data demonstrate that cell growth and suppression of differentiation in rhabdomyosarcoma cells require overproduction of active TGF-beta; furthermore, they show that a 'critical' concentration of TGF-beta is necessary for myogenic differentiation to occur, whereas myogenesis is abolished below and above this concentration. By impairing the TGF-beta autocrine loop, TPA stabilizes the factor concentration within the range compatible for differentiation to occur. In contrast, in human primary muscle cells a much higher concentration of exogenous TGF-beta is required for the differentiation inhibitory effect and TPA inhibits differentiation in these cells probably through a TGF-beta independent mechanism. These data thus clarify the mechanism underlying the multiple roles of TGF-beta in the regulation of both the transformed and differentiated phenotype.

Isolation and characterization of the murine zinc finger coding gene, ZT2: expression in normal and transformed myogenic cells.

In the context of a project aimed at the identification of zinc finger proteins involved in skeletal muscle histogenesis and differentiation, we isolated a murine gene, named ZT2. The 2.44kb partial cDNA clone corresponds to the 3' region of the gene, and contains a 0.54kb open reading frame encoding four C2H2-like zinc finger domains, organized in tandem. This cDNA hybridizes with multiple transcripts (2, 4.5 and 7kb), whose expression levels vary in different tissues and at different developmental stages in the same tissue. At least in skeletal muscle we observed differences in the polyadenylation state of the transcripts at different stages of development. Moreover, ZT2 expression is correlated with cell proliferation and transformation. Sequence analysis and genetic mapping indicate that ZT2 is the homologue of ZNF125, one of the linked zinc finger encoding genes localized on human Chr 11q23. In humans, a high frequency of tumor-associated translocations is found in this chromosome region. As expected, ZT2 maps to the corresponding region on chromosome 9 in the mouse.

Fibrinogen is an independent marker for thoracic aortic atherosclerosis.

The fibrinogen level is an independent risk factor for coronary events and stroke, but no detailed data are available concerning fibrinogen and atherosclerotic disease of the thoracic aorta. This prospective study using multiplane transesophageal echocardiography examined the relation between atherosclerotic thoracic aortic plaque and fibrinogen level. One-hundred forty-eight patients (65 +/- 11 years) with valvular heart disease underwent multiplane transesophageal echocardiography and coronary angiography. We measured plasma fibrinogen level for each patient and recorded the following cardiovascular risk factors: age, sex, systemic hypertension, history of smoking, hypercholesterolemia, diabetes mellitus, body mass index, and family history of coronary artery disease (CAD). Patients with thoracic aortic plaque had a higher level of plasma fibrinogen (p = 0.0001), were older (p = 0.0001), and had significantly more risk factors: history of smoking (p = 0.009), hypertension (p = 0.008), hypercholesterolemia (p = 0.0001), diabetes mellitus (p = 0.01), and family history of CAD (p = 0.003). Multivariate logistic regression analysis of fibrinogen level and risk factors revealed 4 independent predictors of thoracic aortic plaque: fibrinogen, age, hypercholesterolemia, and history of smoking. Fibrinogen was also an independent predictor of CAD. There was a relation between fibrinogen levels and the severity of aortic atherosclerosis (r = 0.46; p = 0.0001) and the severity of CAD (r = 0.30; p = 0.0001). This prospective study indicates that fibrinogen is an independent marker for thoracic aortic plaque related to the severity of thoracic aortic atherosclerosis and confirms that fibrinogen constitutes an independent marker for CAD related to the severity of angiographically evaluated coronary atherosclerosis.

The risk of coronary artery disease after heart transplantation is increased in patients receiving low-dose cyclosporine, regardless of blood cyclosporine levels.

BACKGROUND: Coronary artery disease (CAD) of allografted hearts is the main cause of late mortality after cardiac transplant, but its etiology is still undetermined. HYPOTHESIS: This study was undertaken to evaluate the relevance of several risk factors, including cyclosporine (CsA) dose and blood CsA levels, to the incidence of CAD. METHODS: In 163 heart transplants performed between November 1985 and August 1994 at our Institution, CAD was diagnosed by coronary angiography or at postmortem examination. Patients in whom postmortem examination or coronary angiography was not performed, as well as those < 15 years of age and those who died within 1 month of surgery, were excluded from the study. The following risk factors were analyzed: recipient age, gender, pretransplant diagnosis, donor age, number of human leukocyte antigen (HLA)-AB mismatches, cytomegalovirus serology, mear serum cholesterol and triglyceride levels, the number of treated acute rejections, mean weighted CsA dose (CsA dosew and weighted blood CsA levels (blood CsA levelw). RESULTS: Coronary artery disease was diagnosed in 32 patients (19.6%). A low mean CsA dosew was the only significant predictor for CAD at multivariate analysis (p < 0.01): there was no correlation with blood CsA levelw. In the patients receiving a CsA dosew > 4 mg/kg/day, the 8.9 year probability of their remaining CAD free was 69% [confidence interval (CI) 50-87%] in comparison with 31% (CI 0-65%) in patients receiving a CsA dosew < 4 mg/kg/day. CONCLUSION: In our experience, a low CsA maintenance dose is the main risk factor for CAD, irrespective of blood CsA levels.

Preliminary results of intrathymic injection of donor cells to prevent acute rejection in human heart transplantation.

In rodents, the intrathymic injection of donor cells or major histocompatibility complex peptides induces indefinite survival of a subsequent allograft with little or no immunosuppression. Here, experiments have been performed in two patients with cardiac transplantation to establish (1) the safety and tolerability of the intrathymic injection of donor leukocytes at the time of transplant surgery and (2) whether conventional immunosuppression interfered with the process of the thymic recognition of alloantigens. It was shown that the intrathymic inoculation of donor cells is safe and can be done without undesired effects. However, the procedure, as performed, did not protect from acute graft rejection. There are data enough to attribute the failure of the thymus technique in these two patients to the concomitant use of immunosuppressants. The results of this study are relevant for future trials aimed at finding the appropriate experimental conditions for the use of the thymic approach in human organ transplantation.

Reduction of haemorrhagic complications during mechanically assisted circulation with the use of a multi-system anticoagulation protocol.

Two different anticoagulation protocols were used in 49 consecutive patients mechanically supported either for bridge to transplantation (11) or for recovery of myocardial function after cardiac surgery (35). In 46 patients a Biomedicus centrifugal pump was used and in 3 patients a Pierce-Donachy ventricles. Mechanical support was provided to the left ventricle in 14 patients, to the right ventricle in 6 and to both ventricles in 12 patients; an extra-corporeal membrane oxygenator (ECMO) support was used in 17 patients. Thirty-seven males and 12 females, aged 0.2 to 58 years, were supported for an average time of 6.3 days (range 1-43). Anticoagulation was either based on a continuous infusion of heparin in the first 27 patients (group A) or on a multi-system therapy ("La Pitié" protocol) in the other 22 patients (group B). Overall survival rate was 47%. Patients in group A had a 30% (8/27) survival rate, whereas in group B a 68% (15/22) survival rate was observed (p = 0.006). Transplantation and ventricular assist device (VAD) removal was successfully obtained in 59% (16/27) and 91% (20/22) of patients in group A and group B respectively (p = 0.05). Significant bleeding occurred in 21 patients (81%) in group A and in 2 (9%) of group B (p = 0.001). In these patients bleeding averaged 230 +/- 231 ml/kg in group A versus 55 +/- 18 ml/kg in group B (p = 0.001). Surgical revision was necessary for cardiac tamponade or persistent bleeding in 12 patients of group A (25 procedures: mean 0.9/patient) and in 3 patients of group B (one each patient: mean 0.1/patient) (p = 0.01). Infection, thrombo-embolism and brain hemorrhage were also less frequent in group A than in group B. Our data suggest that the "La Pitié" protocol provides a better control of bleeding than the conventional heparin infusion in patients receiving assist device. this reduction in thrombo-hemorrhagic complications might improve the results of mechanical circulatory support.

Rapid activation and down-regulation of protein kinase C alpha in 12-O-Tetradecanoylphorbol-13-acetate-induced differentiation of human rhabdomyosarcoma cells.

Human rhabdomyosarcoma RD cells express the myogenic regulatory factors MyoD and myogenin but differentiate spontaneously very poorly. Prolonged treatment of RD cells with the protein kinase C (PKC) activator 12-O-tetradecanoylphorbol-13-acetate (TPA) induces growth arrest and myogenic differentiation as shown by the accumulation of alpha-actin and myosin light and heavy chains, without affecting the expression of MyoD and myogenin. In this study, we show that short-term phorbol ester treatment of the cultures is sufficient to trigger myogenic differentiation but not growth arrest. Furthermore, PKC inhibitors, such as staurosporine or calphostin C, prevent TPA-induced differentiation but not cell growth arrest. These data suggest that the two events are mediated by different pathways; a possible interpretation is that the activation of one or more PKC isoforms mediates the induction of differentiation, whereas the down-regulation of the same or different isoforms mediates the growth arrest. To address the mechanism whereby TPA affects cell growth and differentiation in RD cells, we first analyzed PKC isoenzyme distribution. We found that RD cells express the alpha, beta 1, gamma, and sigma PKC isoenzymes. Only the alpha isoform is exclusively found in the soluble fraction, but it translocates to the membrane fraction within 5 min of TPA treatment and is completely down-regulated after 6 h. The other isoenzymes are found associated to both the soluble and the particulate fractions and are down-regulated after long-term TPA treatment. By immunofluorescence analysis, we show that the PKC alpha down-regulation is specific for those cells that respond to TPA by activating the muscle phenotype. We propose that TPA-induced differentiation in RD cells is mediated by the transient activation of PKC alpha, which activates some of the intracellular events that are necessary for MyoD and myogenin transacting activity and for the induction of terminal differentiation of RD cells. By contrast, the constitutively active beta 1 and sigma are responsible for the maintenance of cell growth, and their down-regulation is responsible for long-term TPA-induced cell growth arrest.

Pediatric heart transplantation without chronic maintenance steroids.

From 1986 to February 1993, 40 children aged 2 months to 18 years (average age 10.4 +/- 5.8 years) underwent heart transplantation. Indications for transplantation were idiopathic cardiomyopathy (52%), congenital heart disease (35%) with and without prior repair (71% and 29%, respectively), hypertrophic cardiomyopathy (5%), valvular heart disease (3%), and doxorubicin cardiomyopathy (5%). Patients were managed with cyclosporine and azathioprine. No prophylaxis with antilymphocyte globulin was used. Steroids were given to 39% of patients for refractory rejection, but weaning was always attempted and generally successful (64%). Five patients (14%) received maintenance steroids. Four patients died in the perioperative period and one died 4 months later. There have been no deaths related to rejection or infection. Average follow-up was 36 +/- 19 months (range 1 to 65 months). Cumulative survival is 88% at 5 years. In patients less than 7 years of age, rejection was monitored noninvasively. In the first postoperative month, 89% of patients were treated for rejection. Freedom from serious infections was 83% at 1 month and 65% at 1 year. Cytomegalovirus infections were treated successfully with ganciclovir in 11 patients. No impairment of growth was observed in children who underwent transplantation compared with a control population. Twenty-one patients (60%) have undergone annual catheterizations and no sign of graft atherosclerosis has been observed. Seizures occurred in five patients (14%) and hypertension was treated in 10 patients (28%). No patient was disabled and no lymphoproliferative disorder was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

TPA-induced differentiation of human rhabdomyosarcoma cells: expression of the myogenic regulatory factors.

RD cells (a cell line derived from a human rhabdomyosarcoma) undergo a very limited myogenic differentiation despite the fact that they express several myogenic determination genes. Since we have previously shown (Aguanno et al., Cancer Res. 50, 3377, 1990) that the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induces myogenic differentiation in these cells, in this paper we investigate the mechanism by which TPA interferes with the expression and/or function of the myogenic determination genes. Northern blot analysis revealed that RD cells express the myf3 (the human analog of MyoD) and myf4 (the human analog of myogenin) transcripts, but not myf5 or myf6 transcripts. The myf3 and the myf4 gene products are correctly translated and accumulated in the nuclei as shown by immunofluorescence analysis. The tumor promoter (TPA) does not modify the pattern of expression of the myf factors while it induces the accumulation of muscle-specific transcripts, such as alpha-actin and fast myosin light chain 1, and their corresponding proteins. On the other hand, within 1 day of treatment, TPA inhibits the expression of the Id gene, which is a negative regulator of MyoD activity. However, while the TPA-induced inhibition of Id message accumulation correlates with differentiation, cell confluence also causes a reduction in Id message accumulation, without inducing differentiation. Under our experimental conditions, overexpression of any of the myf cDNAs in RD cells does induce spontaneous differentiation but enhances the effect of TPA treatment independently from the level of the expressed message. These data suggest that differentiation of RD cells is likely to depend upon the activity of complexes containing the various members of the MyoD family, which can be regulated by proteins affecting MyoD dimerization such as Id, but also by other mechanisms induced by TPA, such as phosphorylation.

[Emergency surgery of acute coronary insufficiency]. / La chirurgia d'urgenza nell'insufficienza coronarica acuta.

In our centre, during the last five years, emergency operations (within 6 hours) and urgent operations (within 72 hours) have represented 1/4 of all coronary surgery. 295 patients (pts) have been operated on since 1972: of these, 279 with simple revascularization, 5 with combined major surgery, and 11 as a consequence of mechanical complications of acute myocardial infarction. These last were all in cardiogenic shock: the overall 30-day mortality rate was 5.4% (3.6% in those pts with simple revascularization, 20% in those with combined major surgery, and 45.4% in pts with cardiogenic shock). In the subgroup with simple revascularization, the incidence of non fatal perioperative acute myocardial infarction (AMI) was 4.7% in 253 pts with unstable angina, 52.2% in 23 pts with abrupt closure during coronary angioplasty, and obviously 100% in 3 pts surgically treated during evolving AMI. We were able to identify in the univariate analysis as the only 30-day risk mortality factors: 1) a reduced ejection fraction (< 30%) and 2) the combination with endarterectomy. Other factors (female sex, age > 70, severity of angina, diffuse coronary artery disease and more than 3 by pass grafts) have shown a tendency to increase the mortality rate without statistical significance. No deaths occurred in pts revascularized in emergency situations due to coronary angioplasty complications. In recent years emergency and urgent coronary surgical operations have been increasing, with an increase in pts with higher risk factors. In pts with simple revascularization, 30-day mortality and incidence of myocardial infarction are similar to those of elective surgery. In pts with abrupt closure as a consequence of coronary angioplasty the mortality rate seems very low, while the incidence of infarction remains extremely high. These observations have allowed the development of an integrated protocol of intervention in acute unstable coronary syndromes.

Altered protein phosphorylation in murine muscular dystrophy.

Protein phosphorylation has been studied in the dydy murine muscular dystrophy, both in intact muscle cells and in various membrane fractions derived from them. The results obtained showed that several polypeptides were more heavily phosphorylated in dystrophic myotubes in culture as well as in dystrophic muscle fibers isolated from tibialis anterior. In vitro phosphorylation studies revealed that a large polypeptide of apparent molecular weight of 170,000-150,000 was phosphorylated under basal conditions (3 mM EGTA) in dydy microsomal membranes. The phosphorylation of this polypeptide was not stimulated further by cAMP, calmodulin, cGMP or 12-O-tetradecanoylphorbol 13-acetate (TPA). Under no condition was the corresponding polypeptide phosphorylated at an appreciable rate in normal microsomal membranes. An antibody raised against the voltage-dependent calcium channel reacted, in an immunoblot assay, with a polypeptide, present in both normal and dydy microsomes, which had migration characteristics identical to the phosphorylated 170-150 kDa polypeptide after one- or two-dimensional gel electrophoresis. Additional differences were identified in the phosphorylation of smaller polypeptides of microsomal membranes. When sarcolemmal membranes of normal and dydy muscle were phosphorylated in vitro, no major differences were observed. These results show the existence of an alteration of protein phosphorylation in dystrophic muscle cells in vitro and in vivo, leading to abnormal phosphorylation of the voltage-dependent calcium channel. The possible causes and consequences of this alteration are discussed.

Adrenocorticotropin is a specific mitogen for mammalian myogenic cells.

Peptides derived from proopiomelanocortin (POMC) have been found to stimulate the proliferation of murine myogenic cells. Among these peptides, adrenocorticotropin (ACTH) and alpha-, beta-, and gamma-melanocyte-stimulating hormones (MSH) were found to be active, whereas the opioid peptides were not. At clonal density, both ACTH and MSH caused a three- to fourfold increase in the average number of cells per clone in myogenic but not in fibroblast colonies. At high cell density, ACTH and MSH caused a three- to fourfold increase in proliferation of myogenic cells, reflected by an increased accumulation of skeletal myosin. On the other hand mouse embryo skin or muscle fibroblasts or vertebral chondroblasts did not increase proliferation in response to POMC-derived peptides. The half-maximal dose at which ACTH stimulated myoblast proliferation was around 5 nM, and the mitogenic effect was doubled by suboptimal doses of fibroblast growth factor. The possible physiological significance of the mitogenic effect of ACTH on myogenic cells is discussed.

'Early' mammalian myoblasts are resistant to phorbol ester-induced block of differentiation.

Mesenchymal cells were isolated from somites and limbs of mouse embryos at different developmental stages. When grown in tissue culture, some of the cells underwent muscle differentiation as indicated by synthesis of sarcomeric myosin, acetylcholine receptor and, in the case of limb cells, fusion into multinucleated myotubes. When the tumour promoter 12-O-tetradecanoyl phorbol 13-acetate (TPA) was added to these cultures, it caused differential effects, depending upon the age of the embryo from which cells were isolated. In cultures of somites or limb bud from embryos up to 12 days post coitum, TPA did not interfere with the appearance of differentiated muscle cells. When TPA was added to cultures from older embryos, it inhibited muscle differentiation with an efficiency which increased with the age of the embryo, reaching about 90% inhibition at 15 days. After this period, a new population of myogenic cells appeared in the limb, which were able to differentiate in the presence of TPA and represented the great majority of myoblasts after day 18 of embryonic development. The simplest interpretation of these data can be based on the existence of three major classes of myogenic cell precursors, which appear sequentially during muscle histogenesis: 'early' myoblasts, which appear resistant to tumour promoters; 'late' myoblasts, whose differentiation is inhibited by tumour promoters and 'satellite' cells which, like early myoblasts, show no sensitivity to TPA.

Black widow spider toxin-induced calcium fluxes and transmitter release in a neurosecretory cell line.

Several polypeptide neurotoxins affect presynaptic functions by interfering with chemical neurotransmission. This group of toxins includes botulinum toxin, tetanus toxin, beta-bungaro-toxin and black widow spider toxin (BWSTx). While the effect of the first three toxins is mainly a rapid and severe block of neurotransmitter release, BWSTx affects transmission by a massive stimulation of mediator release. Despite various hypotheses put forward to explain the action of BWSTx at the level of nerve terminals, there is still a considerable degree of uncertainty as to the cation dependence of venom action. Study of the toxin mode of action at the biochamical level has been hampered by the complexity and cellular heterogeneity of the preparations used, neuromuscular junction or synaptosomes. PC12 cell line, derived from a rat phaeochromocytoma, seems to be an excellent model in view of its property of synthesising and storing noradrenaline, dopamine and acetylcholine, and releasing them in depolarising conditions. We have recently shown that highly purified BWSTx stimulates secretion from PC12 cells of previously taken up radioactive dopamine (DA) and noradrenaline (NA) (ref. 14 and manuscript in preparation). We report here that the earliest detectable event after toxin treatment of such cells is a massive increase of cytosolic calcium.