Radial artery bypass graft is a feasible and durable conduit for challenging infrainguinal revascularization: 17 years of Melbourne experience.

OBJECTIVES: The superiority of autogenous venous conduits in infrainguinal bypass surgery is well established. In the absence of suitable leg or arm veins the radial artery can be utilized as an alternative autogenous conduit. In contrast to cardiac surgery, experience with the radial artery as a conduit for infrainguinal bypass surgery is limited. The purpose of this study was to review the outcomes of our radial artery bypasses over the last 17 years. METHODS: All radial artery bypasses performed between 1995 and 2012 were identified from a prospective database. Patency, limb salvage, and survival were calculated using the Kaplan-Meier survival estimate method. RESULTS: Twenty-nine radial artery bypasses were performed in 28 patients. Median follow-up was 55 months (range 1-170). Twelve-month primary, assisted primary, and secondary patency rates were 49%, 62%, and 73% respectively; Both 3-year and 5-year primary, assisted primary, and secondary patency rates were 49%, 56% and 67% respectively. Limb salvage rate was 75% at 1- and 5-year follow-up. Patient survival at 1, 3, and 5 years was 96%, 88%, and 76%. CONCLUSIONS: For patients with need of challenging infrainguinal revascularization without suitable autogenous venous conduit, a radial artery bypass can be performed safely with favorable long-term patency and limb salvage rates.

Prostaglandin production by rheumatoid synovial cells: stimulation by a factor from human mononuclear Cells.

Human peripheral blood mononuclear cells (lymphocyte-monocyte) in culture release a solube factor which can stimulate, up to 200-fold, production of prostaglandin E2 by isolated, adherent, rheumatoid synovial cells. Production of the factor by the mononuclear cells is enhanced by phytohemagglutinin. This factor is similar in apparent mol vt (10,000-20,000) to that which also stimulates collagenase production by the same cells.

Antihemostatic, antiinflammatory, and immunosuppressive properties of the saliva of a tick, Ixodes dammini.

Pilocarpine-induced saliva of the tick, Ixodes dammini, inhibited platelet aggregation triggered by ADP and collagen, as well as platelet-aggregation factor. In addition, we found apyrase activity (which degrades ATP and ADP to AMP and orthophosphate) and an anticoagulant. We showed the presence of prostaglandin E2 (PGE2) by bioassay and radioimmunoassay. This saliva inhibited interleukin 2 production by T cell hybridomas, an activity consistent with that of PGE2. A kininase was demonstrated, and this may counteract the algesia- and edema-promoting properties of PGE2. Together, these salivary components produce antihemostatic, antiinflammatory, and immunosuppressive effects that may facilitate feeding, as well as transmission of tick-borne pathogens.

Cyclooxygenase blockade elevates leukotriene E4 production during acute anaphylaxis in sheep.

We examined changes in the levels of eicosanoids in blood and pulmonary lymph of anesthetized sheep undergoing acute anaphylaxis. Within 1-3 min of intravenous antigenic challenge of previously sensitized sheep, there were approximately 7-30-fold elevations in mean arterial plasma levels of thromboxane B2 and 6-ketoprostaglandin F1 alpha, respectively, as measured by RIA. Negligible changes in levels of these cyclooxygenase products were found in both nonsensitized sheep and in sensitized sheep treated with indomethacin before antigenic challenge. In contrast, no changes in levels of sulfidopeptide leukotrienes (SPLT) in pulmonary lymph were detectable by RIA during anaphylaxis in sensitized or nonsensitized sheep, but levels of SPLT in indomethacin-treated sensitized sheep increased more than fivefold above levels in lymph from both other groups of animals. The immunoreactive SPLT in lymph from indomethacin-treated sheep was accounted for as LTE4, as demonstrated by mobility on HPLC and absorbance at 280 nm. These results support the possibility that certain undesirable effects of nonsteroidal antiinflammatory drugs, such as cardiopulmonary reactions in aspirin-sensitive individuals, and impaired renal and cardiac function during therapy with these drugs, may be related in part to augmented synthesis of the 5-lipoxygenase pathway products, especially those of the sulfidopeptide class. Increased LT production could also limit the antiinflammatory effectiveness of these drugs in many disease states.

The configuration of DNA replication sites within the Trypanosoma brucei kinetoplast.

The kinetoplast is a concatenated network of circular DNA molecules found in the mitochondrion of many trypanosomes. This mass of DNA is replicated in a discrete "S" phase in the cell cycle. We have tracked the incorporation of the thymidine analogue 5-bromodeoxyuridine into newly replicated DNA by immunofluorescence and novel immunogold labeling procedures. This has allowed the detection of particular sites of replicated DNA in the replicating and segregating kinetoplast. These studies provide a new method for observing kinetoplast DNA (kDNA) replication patterns at high resolution. The techniques reveal that initially the pattern of replicated DNA is antipodal and can be detected both on isolated complexes and in replicating kDNA in vivo. In Trypanosoma brucei the opposing edges of replicating kDNA never extend around the complete circumference of the network, as seen in other kinetoplastids. Furthermore, crescent-shaped labeling patterns are formed which give way to labeling of most of the replicating kDNA except the characteristic midzone. The configuration of these sites of replicated DNA molecules is different to previous studies on organisms such as Crithidia fasciculata, suggesting differences in the timing of replication of mini and maxicircles and/or organization of the replicative apparatus in the kinetoplast of the African trypanosome.

Microtubule polarity and dynamics in the control of organelle positioning, segregation, and cytokinesis in the trypanosome cell cycle.

Trypanosoma brucei has a precisely ordered microtubule cytoskeleton whose morphogenesis is central to cell cycle events such as organelle positioning, segregation, mitosis, and cytokinesis. We have defined microtubule polarity and show the + ends of the cortical microtubules to be at the posterior end of the cell. Measurements of organelle positions through the cell cycle reveal a high degree of coordinate movement and a relationship with overall cell extension. Quantitative analysis of the segregation of the replicated mitochondrial genome (the kinetoplast) by the flagellar basal bodies identifies a new G2 cell cycle event marker. The subsequent mitosis then positions one "daughter" nucleus into the gap between the segregated basal bodies/kinetoplasts. The anterior daughter nucleus maintains its position relative to the anterior of the cell, suggesting an effective yet cryptic nuclear positioning mechanism. Inhibition of microtubule dynamics by rhizoxin results in a phenomenon whereby cells, which have segregated their kinetoplasts yet are compromised in mitosis, cleave into a nucleated portion and a flagellated, anucleate, cytoplast. We term these cytoplasts "zoids" and show that they contain the posterior (new) flagellum and associated basal-body/kinetoplast complex. Examination of zoids suggests a role for the flagellum attachment zone (FAZ) in defining the position for the axis of cleavage in trypanosomes. Progression through cytokinesis, (zoid formation) while mitosis is compromised, suggests that the dependency relationships leading to the classical cell cycle check points may be altered in trypanosomes, to take account of the need to segregate two unit genomes (nuclear and mitochondrial) in this cell.

Intramitochondrial localization of universal minicircle sequence-binding protein, a trypanosomatid protein that binds kinetoplast minicircle replication origins.

Kinetoplast DNA (kDNA), the mitochondrial DNA of the trypanosomatid Crithidia fasciculata, is a unique structure containing 5,000 DNA minicircles topologically linked into a massive network. In vivo, the network is condensed into a disk-shaped structure. Replication of minicircles initiates at unique origins that are bound by universal minicircle sequence (UMS)-binding protein (UMSBP), a sequence-specific DNA-binding protein. This protein, encoded by a nuclear gene, localizes within the cell's single mitochondrion. Using immunofluorescence, we found that UMSBP localizes exclusively to two neighboring sites adjacent to the face of the kDNA disk nearest the cell's flagellum. This site is distinct from the two antipodal positions at the perimeter of the disk that is occupied by DNA polymerase beta, topoisomerase II, and a structure-specific endonuclease. Although we found constant steady-state levels of UMSBP mRNA and protein and a constant rate of UMSBP synthesis throughout the cell cycle, immunofluorescence indicated that UMSBP localization within the kinetoplast is not static. The intramitochondrial localization of UMSBP and other kDNA replication enzymes significantly clarifies our understanding of the process of kDNA replication.

Dietary enrichment with the polyunsaturated fatty acid eicosapentaenoic acid prevents proteinuria and prolongs survival in NZB x NZW F1 mice.

Prostaglandins and related compounds are active mediators of inflammation, but data concerning their role in the pathogenesis of the glomerulonephritis of New Zealand Black x New Zealand White (NZB x NZW) F1 mice are conflicting. Dietary eicosapentaenoic acid (EPA, C20:5), a fatty acid analogue of arachidonic acid (C20:4), has been shown to impair platelet aggregation in humans, apparently through inhibition of the synthesis of prostaglandins and thromboxanes from arachidonic acid. We report here the effects of a diet high in EPA on the development of renal disease and survival in female NZB x NZW F1 mice. Animals from 4--5 wk of age were fed diets containing 25% lipid, supplied either as beef tallow or menhaden oil, with fatty acid analysis of less than 0.05 and 14.4% EPA, respectively. In the first experiment, by 13.5 mo of age, mice on the beef tallow diet had all (9/9) developed proteinuria and the majority (6/9) had died, with renal histologic examination revealing severe glomerulonephritis. In contrast, none of 10 menhaden oil-fed animals had developed proteinuria, and all were alive at this time (P less than 0.005 for both proteinuria and survival). In a second experiment using 50 mice in each dietary group, 56% of the beef tallow group vs. none of the menhaden oil group had developed proteinuria at 9 mo of age (P less than 0.005). Native DNA binding at 6 mo of age was 23.9 +/- 14.7 vs. 10.1 +/- 9.7% in the beef and menhaden oil groups, respectively (P less than 0.01). Weights were similar in all groups, and there was no evidence of essential fatty acid deficiency in any group. These results demonstrate that a diet high in EPA protects NZB x NZW F1 mice from the development of glomerulonephritis.

Prostaglandin-stimulated bone resorption by rheumatoid synovia. A possible mechanism for bone destruction in rheumatoid arthritis.

Synovial tissue from patients with rheumatoid arthritis was maintained in organ culture for 3-14 days. Conditioned media from these synovial cultures contained bone resorption-stimulating activity, measured in vitro by using calcium release from mouse calvaria as the assay system. The synovial cultures also produce prostaglandin E2 (PGE2) as measured by serologic methods. The production of both the bone resorption-stimulating activity and PGE2 was inhibited by more than 90% by treatment of the synovial cultures with indomethacin (5 mug/ml). In contrast, treatment of the synovial cultures with colchicine (0.1 mug/ml) caused a marked and parallel increase in the concentration of both bone resorption-stimulating activity and PGE2 in the conditioned media. The bone resorption-stimulating activity was quantitatively extracted into diethyl ether. Within the limits of experimental error, all of the bone resorption-stimulating activity in medium was accounted for by its content of PGE2, itself a potent osteolytic factor. We conclude that the bone resorption-stimulating activity produced by rheumatoid synovia in culture is PGE2.

Dietary menhaden oil lowers plasma prostaglandins and calcium in mice bearing the prostaglandin-producing HSDM1 fibrosarcoma.

The omega 3 class of polyunsaturated fatty acids, particularly eicosapentaenoic acid (EPA, 20:5), has been shown to alter the patterns of arachidonic acid (20:4) metabolism in both in vitro and in vivo systems. To examine further the role of arachidonic acid conversion to prostaglandins (PG) in hypercalcemic mice bearing the PG-producing HSDM1 fibrosarcoma, we have performed experiments in which control and tumor-bearing animals were fed diets either low (0.1-0.2% of total fatty acid) or high (17%) in EPA. In all five experiments performed, tumor-bearing mice eating control diets had markedly elevated (average sixfold above control) plasma concentrations of 13,14-dihydro-15-keto-PGE2 (PGE2-M), while in mice bearing HSDM1 tumors and eating the EPA-enriched menhaden oil diet, the elevation was reduced to only twice control values. The increase in plasma calcium concentration (approximately 2.5 mg/dl above control) in tumor-bearing animals was also reduced significantly (P less than 0.05) to only 1.3 mg/dl above control in mice eating the diet enriched in EPA. Plasma immunoreactive hydroxy fatty acids (i12-HETE) and sulfidopeptide leukotrienes (iSRS) were not elevated in tumor-bearing mice and were unaffected by diet. The contents of PGE2, PGF2 alpha, and 6-keto-PGF1 alpha were lower in tumor tissue from animals eating the diet high in EPA, whereas the tissue contents of i12-HETE and iSRS were not altered by diet. Fatty acid analysis of liver and tumor tissue revealed marked increases in certain omega 3 fatty acids (20:5, 22:5, and 22:6) from animals eating the enriched diet. Body weights, tumor weights, and tumor histology were not significantly altered by diet. To determine whether dietary calcium played a role in the elevation of plasma calcium in mice bearing the HSDM1 tumor and the reduction of plasma calcium in animals fed EPA, we compared results in mice fed diets containing 0.80% (normal) and 0.015% (deficient) calcium. The increases in plasma calcium and PGE2-M observed in tumor-bearing mice were the same on both normal and very low calcium intakes. We conclude, in mice of the Swiss albino strain bearing the HSDM1 fibrosarcoma, that consumption of a diet enriched in EPA reduces the production of cyclooxygenase products of arachidonic acid metabolism and thereby reduces the elevation of plasma calcium concentration. Dietary enrichment with EPA did not alter the production of serologically determined lipoxygenase products of arachidonic acid.

Dietary omega-3 polyunsaturated fatty acids inhibit phosphoinositide formation and chemotaxis in neutrophils.

Earlier studies demonstrated that dietary omega-3 polyunsaturated fatty acid (PUFA) supplementation attenuates the chemotactic response of neutrophils and the generation of leukotriene (LT) B4 by neutrophils stimulated with calcium ionophore; however, the mechanisms and relationship of these effects were not examined. Neutrophils and monocytes from eight healthy individuals were examined before and after 3 and 10 wk of dietary supplementation with 20 g SuperEPA daily, which provides 9.4 g eicosapentaenoic acid (EPA) and 5 g docosahexaenoic acid. The maximal neutrophil chemotactic response to LTB4, assessed in Boyden microchambers, decreased by 69% after 3 wk and by 93% after 10 wk from prediet values. The formation of [3H]inositol tris-phosphate (IP3) by [3H]inositol-labeled neutrophils stimulated by LTB4 decreased by 71% after 3 wk (0.033 +/- 0.013% [3H] release, mean +/- SEM) and by 90% after 10 wk (0.011 +/- 0.011%) from predict values (0.114 +/- 0.030%) as quantitated by beta-scintillation counting after resolution on HPLC. LTB4-stimulated neutrophil chemotaxis and IP3 formation correlated significantly (P < 0.0001); each response correlated closely and negatively with the EPA content of the neutrophil phosphatidylinositol (PI) pool (P = 0.0003 and P = 0.0005, respectively). Neither the affinities and densities of the high and low affinity LTB4 receptors on neutrophils nor LTB4-mediated diglyceride formation changed appreciably during the study. Similar results were observed in neutrophils activated with platelet-activating factor (PAF). The summed formation of LTB4 plus LTB5 was selectively inhibited in calcium ionophore-stimulated neutrophils and was also inhibited in zymosan-stimulated neutrophils. The inhibition of the summed formation of LTB4 plus LTB5 in calcium ionophore-stimulated neutrophils and in zymosan-stimulated neutrophils did not correlate significantly with the EPA content of the PI pool. The data indicate that dietary omega-3 PUFA supplementation inhibits the autoamplification of the neutrophil inflammatory response by decreasing LTB4 formation through the inactivation of the LTA epoxide hydrolase and independently by inhibiting LTB4- (and PAF) stimulated chemotaxis by attenuating the formation of IP3 by the PI-selective phospholipase C. This is the initial demonstration that dietary omega-3 PUFA supplementation can suppress signal transduction at the level of the PI-specific phospholipase C in humans.

The identification of monoclonality in human aberrant crypt foci.

Malignant neoplasms, including colon cancers, are thought to arise from a single initiated progenitor cell. Aberrant crypt foci (ACF) are putative precursors of at least some colon cancers. The pattern of X chromosomal inactivation, which is identified by the differential methylation of a site near a polymorphic CAG repeat in the androgen receptor gene, was used to determine the clonality status of 11 ACF from eight female patients. Ten of 11 ACF were found to be monoclonal aberrations. The eleventh ACF appeared monoclonal, but nonrandom inactivation of the X chromosome was also seen in normal crypts from this patient. These results clearly demonstrate that: (a) a high percentage of ACF lesions are neoplastic rather than hyperplastic; and (b) ACF are the earliest identified neoplastic lesions in the colon.

Predictors of mortality in patients with cardiogenic shock treated with primary percutaneous coronary intervention and intra-aortic balloon counterpulsation.

BACKGROUND: Cardiogenic shock remains the most serious complication of patients hospitalized with acute myocardial infarction (AMI). Early revascularization is the cornerstone of invasive therapy, while mechanical support with intra-aortic balloon pump (IABP) is debatable. From our institutional shock registry we sought to determine predictors of in-hospital mortality-including the aspect of IABP timing-and to develop a clinical risk score for shock patients with AMI. METHODS: From January 2005 till December 2010, 102 patients with cardiogenic shock due to AMI treated with primary percutaneous coronary intervention (PCI) and IABP were analyzed. Univariate and multivariate logistic regression analyses were used to identify independent predictors of in-hospital mortality. Logistic regression analysis and receiver-operating curves were used to generate a mortality risk score. RESULTS: The mean age of the cohort was 70.1 ± 11.0 years and 70 % were men. One third of patients had a non-ST segment elevation myocardial infarction and 30 % had to be resuscitated before coronary intervention. Mean left ventricular ejection fraction was 25 %. After admission, 23 % of patients developed an acute renal failure and 10 % needed renal dialysis during hospital stay. In 52 % of patients IABP therapy was initiated after primary PCI, while the remaining patients had an IABP-assisted primary PCI. All-cause in-hospital mortality was 40.2 %. Using multivariate analysis, age (odds ratio [OR] 1.08, p = 0.006), resuscitation before PCI (OR 3.46, p = 0.045), vasopressor use (OR 7.88, p = 0.003), acute renal failure (OR 11.18, p = 0.001), and IABP implantation after PCI (OR 4.36, p = 0.011) were independently associated with in-hospital mortality. Based on these predictors, a mortality-risk score was calculated as follows: 1.5 × IABP timing before PCI + 0.1 × age + resuscitation before PCI + 2 × vasopressor use + 2.5 × acute renal failure. Using a cut-off value of 10.4, this score had a specificity of 83 % and a sensitivity of 82 % for prediction of in-hospital death. CONCLUSIONS: We identified age, vasopressor use, resuscitation before PCI, acute renal failure and IABP implantation after PCI as independent predictors of in-hospital mortality in patients with cardiogenic shock due to AMI. The timing of IABP insertion was the only modifiable factor predicting in-hospital mortality in our cohort. Consequently, balloon pumping should be started before PCI to improve outcome of cardiogenic shock patients.

The protein tyrosine kinase family of the human genome.

As the sequencing of the human genome is completed by the Human Genome Project, the analysis of this rich source of information will illuminate many areas in medicine and biology. The protein tyrosine kinases are a large multigene family with particular relevance to many human diseases, including cancer. A search of the human genome for tyrosine kinase coding elements identified several novel genes and enabled the creation of a nonredundant catalog of tyrosine kinase genes. Ninety unique kinase genes can be identified in the human genome, along with five pseudogenes. Of the 90 tyrosine kinases, 58 are receptor type, distributed into 20 subfamilies. The 32 nonreceptor tyrosine kinases can be placed in 10 subfamilies. Additionally, mouse orthologs can be identified for nearly all the human tyrosine kinases. The completion of the human tyrosine kinase family tree provides a framework for further advances in biomedical science.

Expiratory flow limitation in large and small airways.

Obstructive airways diseases constitute 75% of all chronic respiratory diseases. This article briefly reviews the physlologic basis for flow limitation in large and small airways. Application of the physiologic principles described can lead to a better understanding of the normal mechanics of breathing and the various alterations resulting from diseases that limit airflow.

Prostaglandins and the mechanism of action of anti-inflammatory drugs.

Aspirin and a large number of nonsteroidal anti-inflammatory drugs act primarily through the inhibition of prostaglandin synthesis by inhibiting the enzyme cyclooxygenase. Other groups of biologically active polyunsaturated fatty acid derivatives including the leukotrienes, are generally not inhibited by this class of drugs in the same concentration ranges. Inhibition of the vasodilator prostaglandins, prostaglandin E2 and prostacyclin, as well as the leukotrienes, may reduce their inflammatory effects in several disease states. In addition, prostaglandin synthesis is also inhibited by glucocorticoids even though their mode of action may involve other effects as well. Prostaglandin E2 stimulates the osteoclastic reabsorption of juxtaarticular bone; its inhibition by nonsteroidal anti-inflammatory agents may, therefore, retard the process of bone erosion in rheumatoid arthritis and in other inflammatory processes. Inhibition of prostaglandin synthesis by these drugs accounts for many of their major toxic effects, including gastritis, which is the most common side effect; precipitation or aggravation of renal failure; fluid retention; hyperkalemia; antiplatelet effects with hemorrhagic phenomena; and aggravation of asthma and rhinosinusitis. Inhibition of prostaglandin synthesis can, therefore, account for most of the therapeutic as well as toxic effects of the nonsteroidal anti-inflammatory agents. Inhibition of pathways of synthesis of other important mediators, such as leukotrienes, are currently under investigation and may provide another approach for the development of new therapeutic agents.

Management of gastrointestinal toxicity of nonsteroidal anti-inflammatory drugs during the therapy of rheumatic diseases. The rheumatologist's perspective.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are the most widely used therapeutic agents for many important rheumatic diseases. A large body of evidence indicates that the anti-inflammatory effects as well as the toxic side effects of NSAIDs can be attributed to their ability to inhibit prostaglandin synthesis. The most frequent side effect leading to discontinuation of NSAID therapy is upper gastrointestinal toxicity. This is a major problem for rheumatologists and their patients since alternatives to NSAIDs are not readily available. The mechanism of upper gastrointestinal toxicity associated with NSAIDs can be attributed to their common ability to inhibit gastric PGE2 synthesis, which suggests that gastrointestinal side effects often may not be avoided by substituting another NSAID, a prediction borne out by many patients with rheumatic diseases. However, there is some evidence that some patients may differ in their susceptibility to gastrointestinal and other toxic effects, though this concept has not been rigorously studied in clinical trials. Thus, although changing the specific NSAID may prevent gastrointestinal toxicity in some patients, many are intolerant to all NSAIDs. Patients who benefit from a specific NSAID, but in whom gastrointestinal toxicity develops, have limited options. Another NSAID may be substituted but at the risk of losing therapeutic efficacy and/or of continuing toxicity. Alternatively, the side effects may be controlled while continuing therapy with the NSAID by one of several measures, including the use of antacids or a histamine (H2)-receptor antagonist, such as cimetidine. The use of enteric-coated salicylates or nonacetylated salicylates may also provide satisfactory alternatives. However, patients who have experienced major gastrointestinal toxicity, such as massive bleeding or active peptic ulcers, should usually avoid NSAID therapy.

Unusual renal manifestations of Wegener's granulomatosis. Report of two cases.

Two patients with previously undescribed renal manifestations of Wegener's granulomatosis are described. A 24 year old man, who presented with typical necrotizing granulomatous sinus disease and cavitary pulmonary lesions, had multiple bilateral renal arterial aneurysms demonstrated angiographically. One of these aneurysms ruptured, leading to a massive perinephric hematoma. The bleeding artery was successfully occluded with Gelfoam embolization, thereby obviating the need for nephrectomy. A 60 year old woman presented with glomerulonephritis and mononeuritis multiplex two years before the development of classic necrotizing granulomatous inflammation of her sinuses and nose, along with pulmonary nodules. In addition, her left ureter became obstructed due to necrotizing vasculitis of the periureteral vessels. Both patients responded dramatically to cyclophosphamide therapy. The diagnosis of Wegener's granulomatosis should be considered in patients who present with multiple renal aneurysms, a spontaneous perinephric hematoma, necrotizing glomerulitis or ureteral obstruction due to vasculitis, even though the characteristic granulomatous respiratory involvement may be absent at that time. It is important to recognize these unusual renal manifestations as features of Wegener's granulomatosis because of the therapeutic efficacy of cytotoxic immunosuppressive agents in this disease.

Characterization and disruption of a new Trypanosoma brucei repetitive flagellum protein, using double-stranded RNA inhibition.

In Trypanosoma brucei, we have cloned a gene approximately 5 kb downstream of the glucose transporter gene cluster, containing a variable number of 102 bp repeats. This gene encodes a protein with no homologues in the data bases. Antibodies raised against the 34 amino acids repeated motif recognized proteins ranging from 145 to 270 kDa, depending on strains, in both bloodstream and procyclic forms of T. brucei. A correlation was established between the apparent molecular mass of the detected proteins and the number of 34 amino acid repeats which varies from 3 to 40. We have called this protein the flagellum transition zone component (FTZC) due to its localization to the proximal region of the axoneme, within the transition zone. FTZC is the only reported example of a trypanosomal protein present in the transition zone. To determine the role of FTZC we developed a new strategy of gene inactivation based on conditional expression of double-stranded RNA. In the presence of tetracycline, expression of the double-stranded RNA, we observed a complete disappearance of FTZC in the EATRO 1125 and EATRO 427 strains of T. hrucei. Molecular ablation of FTZC does not generate any obvious phenotype such as, lethality, modification of growth rate or cellular shape, in the growth conditions used.

Antivimentin antibodies are an independent predictor of transplant-associated coronary artery disease after cardiac transplantation.

BACKGROUND: Transplant-associated coronary artery disease (TxCAD) is the most serious long-term complication after cardiac transplantation. Anti-endothelial antibodies are associated with disease, and one of the major endothelial antigens recognized in the sera of patients has been shown to be the protein filament vimentin. In this study, we investigated whether antivimentin antibodies are associated with TxCAD and whether their presence can be used to identify patients at high risk of developing angiographically detectable TxCAD. METHODS: Up to 5 years after transplantation, 880 sequential sera (7.07+/-1.8 samples/patient) were collected retrospectively from 109 patients; the majority were collected in the first 2 years. Sera were assessed for antivimentin antibodies using ELISA. TxCAD was assessed by annual angiography. RESULTS: Mean titres of antivimentin antibodies, calculated up to 1, 2, and 5 years, were significantly higher in patients who developed TxCAD than those who remained disease free (P<0.0001, P<0.0038, and P<0.0001, respectively). A predictive test based on the first-year mean vimentin titre alone (> or = 120) produced a test with 63% sensitivity and 76% specificity. Inclusion of persistent rejection or high 1-year mean titre (> or = 270) as a risk factor produced a test with 66% sensitivity and 82% specificity. Multivariate analysis of time to occurrence of transplant vasculopathy showed that mean titre at 1 or 2 years was an independent predictor of time until disease in the presence of all other variables. CONCLUSIONS: Antivimentin antibodies are an independent predictor of TxCAD and can be used to identify some of the patients who are at high risk of developing this complication.