Gene therapy for rhesus monkeys heterozygous for LDL receptor deficiency by balloon catheter hepatic delivery of helper-dependent adenoviral vector.

Autosomal dominant familial hypercholesterolemia (FH) is a monogenic life-threatening disease. We tested the efficacy of low-density lipoprotein receptor (LDLR) gene therapy using helper-dependent adenoviral vector (HDAd) in a nonhuman primate model of FH, comparing intravenous injection versus intrahepatic arterial injection in the presence of balloon catheter-based hepatic venous occlusion. Rhesus monkeys heterozygous for mutant LDLR gene (LDLR+/-) developed hypercholesterolemia while on a high-cholesterol diet. We treated them with HDAd-LDLR either by intravenous delivery or by catheter-based intrahepatic artery injection. Intravenous injection of ⩽1.1 × 10(12) viral particles (vp) kg(-1) failed to have any effect on plasma cholesterol. Increasing the dose to 5 × 10(12) vp kg(-1) led to a 59% lowering of the plasma cholesterol that lasted for 30 days before it returned to pre-treatment levels by day 40. A further increase in dose to 8.4 × 10(12) vp kg(-1) resulted in severe lethal toxicity. In contrast, direct hepatic artery injection following catheter-based hepatic venous occlusion enabled the use of a reduced HDAd-LDLR dose of 1 × 10(12) vp kg(-1) that lowered plasma cholesterol within a week, and reached a nadir of 59% pre-treatment level on days 20-48 after injection. Serum alanine aminotransferase remained normal until day 48 when it went up slightly and stayed mildly elevated on day 72 before it returned to normal on day 90. In this monkey, the HDAd-LDLR-induced trough of hypocholesterolemia started trending upward on day 72 and returned to pre-treatment levels on day 120. We measured the LDL apolipoprotein B turnover rate at 10 days before, and again 79 days after, HDAd-LDLR treatment in two monkeys that exhibited a cholesterol-lowering response. HDAd-LDLR therapy increased the LDL fractional catabolic rate by 78 and 50% in the two monkeys, coincident with an increase in hepatic LDLR mRNA expression. In conclusion, HDAd-mediated LDLR gene delivery to the liver using a balloon catheter occlusion procedure is effective in reversing hypercholesterolemia in a nonhuman primate FH model; however, the unsustainability of the hypocholesterolemic response during 3-4 months of follow up and heterogeneous response to the treatment remains a challenge.

Long-term follow-up of the STARFlex device for closure of secundum atrial septal defect.

BACKGROUND: There is limited published outcome data on the STARFlex device for transcatheter closure of atrial septal defects (ASD). AIM: To contribute to the long term outcomes of ASD device closure with the STARFlex device. METHODS: Results of a prospective FDA approved clinical trial (1999-2001) from a single institution of the STARFlex device for simple ASD was reviewed. The inclusion criteria included age >or=2 years, isolated secundum ASD, evidence of right ventricular volume overload, and maximum stretched diameter <22 mm. A retrospective review of the original study data with most recent follow-up was performed. Clinical success in the trial was defined as complete closure or residual leak

Assessing the importance of genotype x environment interaction for root traits in rice using a mapping population II: conventional QTL analysis.

Modifying plant root systems is considered a means of crop improvement targeted to low-resource environments, particularly low nutrient and drought-prone agriculture. The identification of quantitative trait loci (QTLs) for root traits has stimulated marker-assisted breeding to this end, but different QTLs have been detected in different populations of the same species, and importantly, in the same population when grown in different experimental environments. The presence of QTL x environment interaction is implicated, and this must be characterised if the utility of the target QTLs is to be realised. Previous attempts to do this suffer from a lack of control over replicate environments and inadequate statistical rigour. The Bala x Azucena mapping population was grown in two replicate experiments of four treatment environments, a control, a low light, a low soil nitrogen and a low soil water treatment. After a 4 weeks growth, maximum root length, maximum root thickness, root mass below 50 cm, total plant dry mass, % root mass and shoot length were measured. A summary of the overall results is presented in an accompanying paper. Here, QTL analysis by composite interval mapping is presented. A total of 145 QTLs were detected, mapping to 37 discrete loci on all chromosomes. Superficial evidence of QTL x E (great difference in LOD score) was tested by single-marker analysis which confirmed QTL x E for five loci representing only five individual trait-loci interactions. Some loci appeared to be stable across environments. Some QTLs were clearly more or less active under low light, low nitrogen or drought. A few notable loci on chromosomes 1, 2, 3, 5, 7 and 9 are briefly discussed. Also discussed are some remaining statistical shortcomings that will be addressed in another companion paper.

Influence of nematodes on resource utilization by bacteria--an in vitro study.

The positive influence of bacterial feeding nematodes on bacterial mediated processes such as organic matter mineralization and nutrient cycling is widely accepted, but the mechanisms of these interactions are not always apparent. Both transport of bacteria by nematodes, and nutritional effects caused by nematode N excretion are thought to be involved, but their relative importance is not known because of the difficulties in studying these interactions in soil. We developed a simple in vitro assay to study complex nematode/bacterial interactions and used it to conduct a series of experiments to determine the potential influence of nematode movement and nutritional effects on bacterial resource use. The system used bacterial feeding and nonfeeding insect parasitic nematodes, and luminescent bacteria marked with metabolic reporter genes. Both nutritional enhancement of bacterial activity and bacterial transport were observed and we hypothesize that in nature, the relative importance of transport is likely to be greater in bulk soil, whereas nematode excretion may have greater impact in the rhizosphere. In both cases, the ability of nematodes to enhance bacterial resource utilization has implications for soil components of biogeochemical cycling.

Assessing the importance of genotype x environment interaction for root traits in rice using a mapping population. I: a soil-filled box screen.

Altering root system architecture is considered a method of improving crop water and soil nutrient capture. The analysis of quantitative trait loci (QTLs) for root traits has revealed inconsistency in the same population evaluated in different environments. It must be clarified if this is due to genotype x environment interaction or considerations of statistics if the value of QTLs for marker-assisted breeding is to be estimated. A modified split-plot design was used where a main plot corresponded to a separate experiment. The main plot factor had four treatments (environments), which were completely randomized among eight trials, so that each treatment was replicated twice. The sub-plot factor consisted of 168 recombinant inbreed lines of the Bala x Azucena rice mapping population, randomly allocated to the seven soil-filled boxes. The aim of the trial was to quantify QTL x environment interaction. The treatments were chosen to alter partitioning to roots; consisting of a control treatment (high-soil nitrogen, high light and high-water content) and further treatments where light, soil nitrogen or soil water was reduced singly. After 4 weeks growth, maximum root length (MRL), maximum root thickness, root mass below 50 cm, total plant dry mass (%), root mass and shoot length were measured. The treatments affected plant growth as predicted; low nitrogen and drought increased relative root partitioning, low-light decreased it. The parental varieties Bala and Azucena differed significantly for all traits. Broad-sense heritability of most traits was high (57-86%). Variation due to treatment was the most important influence on the variance, while genotype was next. Genotype x environment interaction was detected for all traits except MRL, although the proportion of variation due to this interaction was generally small. It is concluded that genotype x environment interaction is present but less important than genotypic variation. A companion paper presents QTL x environment analysis of data.

Patent foramen ovale morphology and impact on percutaneous device closure.

We sought to evaluate the effects of atrial septal and patent foramen ovale (PFO) morphology on the efficacy of transcatheter closure. We performed a retrospective analysis of all patients who underwent PFO device closure from January 1997 to January 2002. Forty-seven patients underwent percutaneous closure of a PFO with a median age of 45 years (range, 8-75) and weight of 76 kg (range 28-115). The septal morphology was flat in 33 and aneurysmal in 14 patients. The PFO morphology was a simple flap in 20 and complex in 27 patients. Complex morphologies included long-tunnel PFO (n = 15), coexistent small atrial septal defect (ASD) (n = 5), and aneurysmal septum without a tunnel or ASD (n = 7). Nonstretched PFO diameters were significantly smaller than stretched (4.8 +/- 1.1 mm vs 11.6 +/- 3.8 mm, p < 0.01). Median device size: stretched diameter ratio (DS:SD) was 3.7:1 (range, 2.2-9.1). The DS:SD ratio was significantly higher in patients with complex PFO (mean, 3.9:1 vs 2.6:1; p < 0.05). Device placement was successful in all patients. Five patients required transeptal puncture of the foraminal flap in long-tunnel PFOs. Effective closure on follow-up was achieved in 45 patients (95%). Of the 2 patients with residual shunts, 1 had a complex PFO and the other a simple PFO. Two patients (5%) experienced recurrent neurologic symptoms after device closure despite having no residual shunt by echocardiography. Complex PFO did not increase risk of residual shunt or recurrent neurologic symptoms after device closure.

Influence of Soil Temperature and Matric Potential on Sugar Beet Seedling Colonization and Suppression of Pythium Damping-Off by the Antagonistic Bacteria Pseudomonas fluorescens and Bacillus subtilis.

ABSTRACT Pseudomonas fluorescens B5 and Bacillus subtilis MBI 600 colonized sugar beet seedlings at matric potentials of -7 x 10(3), -140 x 10(3), and -330 x 10(3) Pa and under five temperature regimes ranging from 7 to 35 degrees C, with diurnal fluctuations of 5 to 22 degrees C. No interaction between matric potential and temperature was observed. In situ bioluminescence indicated physiological activity of Pseudomonas fluorescens B5. Colonization of the root at >/=4 cm below the seed decreased at very low matric potential (-330 x 10(3) Pa). Total population size of Pseudomonas fluorescens B5 per seedling was significantly increased at -140 x 10(3) Pa. However, matric potential had no significant effect on the population density of Pseudomonas fluorescens per gram of root fresh weight and did not affect the distribution of the population down the root. Total population size per seedling and downward colonization by Pseudomonas fluorescens B5 were significantly reduced at high temperatures (25 to 35 degrees C). Maximum colonization down the root occurred at intermediate temperature (15 degrees C) at both matric potentials (-7 x 10(3) and -140 x 10(3) Pa). Addition of B. subtilis MBI 600 to the seed had no effect on rhizosphere populations of Pseudomonas fluorescens B5. Populations of B. subtilis MBI 600, which consisted largely of spores, were slightly reduced at lower matric potentials and were not affected by temperature. Survival and dry weight of plants in soils infested with Pythium spp. decreased with increasing soil temperature and matric potential, indicating an increase in disease pressure. However, there was no significant interaction between the two factors. At -330 x 10(3) Pa, soil dryness but not Pythium infection was the limiting factor for plant emergence. At temperatures of 7 to 25 degrees C and matric potentials of -7 x 10(3) to 120 x 10(3) Pa, treatment with Pseudomonas fluorescens B5 increased plant survival and dry weight. At 7 degrees C and -120 x 10(3) Pa, there was almost complete emergence of seeds treated with Pseudomonas fluorescens B5. Antagonistic activity of Pseudomonas fluorescens B5 decreased with increasing soil temperature and decreasing matric potential. At 25 to 35 degrees C and -7 x 10(3) Pa, no effect was observed. In regimes with different day and night temperatures, the maximum (day) temperature was decisive for disease development and antagonistic activity. B. subtilis MBI 600 displayed no significant antagonistic effect against Pythium ultimum and did not influence the performance of Pseudomonas fluorescens B5 in combined inocula.

Nematode-enhanced microbial colonization of the wheat rhizosphere.

The mechanisms by which seed-applied bacteria colonize the rhizosphere in the absence of percolating water are poorly understood. Without mass flow, transport of bacteria by growing roots or soil animals, particularly nematodes may be important. We used a sand-based microcosm system to investigate the ability of three species of nematodes (Caenorhabditis elegans, Acrobeloides thornei and a Cruznema sp.) to promote rhizosphere colonization by four strains of beneficial rhizobacteria. In nearly all cases, rhizosphere colonization was substantially increased by the presence of nematodes, irrespective of bacterial or nematode species. Our results suggest that nematodes are important vectors for bacteria rhizosphere colonization in the absence of percolating water.

Intrastent sonotherapy in pulmonary vein restenosis: a new treatment for a recalcitrant problem.

A 2 year old boy developed recurrent pulmonary vein stenosis after surgical repair of infradiaphragmatic pulmonary venous connection. He had required implantation of stents in the left and right sided pulmonary veins at 7 and 13 months of age, respectively. By 2 years of age he had undergone three catheterisation procedures and two surgical procedures to treat recurrent pulmonary vein stenosis. His right ventricular pressure was suprasystemic and catheterisation showed severe neointimal proliferation of both left and right sided stents. At this time the stents were dilated by balloon with simultaneous intrastent sonotherapy. Three months later the patient's clinical improvement was significant, his right ventricular pressure had decreased, and Doppler velocity had decreased across both left and right sided stents.

Use of the transseptal puncture in transcatheter closure of long tunnel-type patent foramen ovale.

Two patients with long tunnel-type patent foramen ovale presented for elective transcatheter closure following transient ischaemic attack and stroke. Right to left shunting was confirmed on transthoracic and transoesophageal echocardiography. A new technique that used a transseptal procedure was devised to enable closure of the tunnel-type patent foramen ovale using the CardioSEAL transseptal occluder to avoid "bunching up" of the device and residual transatrial shunting.

Sizing of atrial septal defects to predict successful closure with transcatheter cardioSEAL device.

We conducted this retrospective study to compare methods for measuring atrial septal defects and to identify factors affecting echocardiographic measurement of such defects before transcatheter closure with the CardioSEAL'Septal Occluder. We reviewed the records of patients considered for device placement at our institution from January 1997 to April 1999. Atrial septal defect size was measured by transthoracic and transesophageal echocardiography; the stretched diameter was measured during catheterization by fluoroscopy and transesophageal echocardiography. The stretched-diameter fluoroscopic measurement was used for device size selection. Analysis of variance was used to calculate the effect of size, age, and size-by-age interaction. Thirty-one patients (3.3 to 72 years of age) underwent transthoracic and transesophageal echocardiography One patient was excluded from catheterization because of a 25-mm septal defect as indicated by transesophageal echocardiography (our maximum diameter, 15 mm). Thirty patients underwent transcatheter stretched-diameter sizing; 5 were excluded from device implantation because of defects >20 mm by stretched-diameter fluoroscopy (4) or septal length insufficient for device support (1). Implantation was successful in 23/25 patients; 2/23 had a residual shunt. In patients with available results (26/30), the stretched diameter was the same whether measured by stretched-diameter fluoroscopy or transesophageal echocardiography (P=0.007 R square=0.963). Compared with stretched-diameter fluoroscopy, precatheterization transthoracic and transesophageal echocardiography underestimated defect size by a mean of 22% and 13.2%, respectively. When data from those same tests were compared in defects of < or =0 mm and > 10 mm, transthoracic and transesophageal echocardiography were reliable predictors (P=0.003 and P=0.05, respectively) of stretched-diameter size in defects < or =0 mm.

Safety of magnetic resonance imaging immediately following Palmaz stent implant: a report of three cases.

Magnetic resonance imaging (MRI) can provide important information on patients with congenital heart defects. There is some reluctance to perform MRI acutely following intravascular stent implant, due to concerns of distortion or movement of the stent in the magnetic field. We report on three patients who underwent MRI evaluation less than 14 days following Palmaz stent implant in the pulmonary arteries and superior vena cava, with no acute adverse outcome or long-term problems.

Redilation of endovascular stents in congenital heart disease: factors implicated in the development of restenosis and neointimal proliferation.

OBJECTIVES: We sought to determine the incidence of and risk factors for the development of restenosis and neointimal proliferation after endovascular stent implantation for congenital heart disease (CHD). BACKGROUND: Risk factors for the development of restenosis and neointimal proliferation are poorly understood. METHODS: This was a retrospective review of patients who underwent endovascular stent redilation between September 1989 and February 2000. RESULTS: Of 368 patients who had 752 stents implanted, 220 were recatheterized. Of those 220 patients, 103 underwent stent redilation. Patients were classified into three groups: 1) those with pulmonary artery stenosis (n = 94), tetralogy of Fallot/pulmonary atresia (n = 72), congenital branch pulmonary stenosis (n = 9), status post-Fontan operation (n = 6), status post-arterial switch operation (n = 7); 2) those with iliofemoral venous obstruction (n = 6); and 3) those with miscellaneous disorders (n = 3). The patients' median age was 9.9 years (range 0.5 to 39.8); their mean follow-up duration was 3.8 years (range 0.1 to 10). Indications for stent redilation included somatic growth (n = 67), serial dilation (n = 27) and development of neointimal proliferation or restenosis, or both (n = 9). There was a low incidence of neointimal proliferation (1.8%) and restenosis (2%). There were no deaths. Complications included pulmonary edema (n = 1), hemoptysis (n = 1) and contralateral stent compression (n = 2). CONCLUSIONS: Redilation or further dilation of endovascular stents for CHD is effective as late as 10 years. The risk of neointimal proliferation (1.8%) and restenosis (2%) is low and possibly avoidable. Awareness of specific risk factors and modification of the stent implantation technique, including avoidance of minimal stent overlap and sharp angulation of the stent to the vessel wall and avoidance of overdilation, have helped to reduce the incidence of restenosis.

Three new applications of stent technology in a single patient.

The use of endovascular stents has become widely established in maintaining both arterial and venous patency in congenital heart disease. Stent implantation is now applied to pulmonary arterial stenoses, coarctation, pulmonary and systemic venous obstruction, and obstructed homografts and conduits, in both the pediatric and adult populations. The purpose of this report is to describe 3 new applications of stent technology: 1) double pulmonary artery stent implantation with simultaneous balloon dilation of a previously placed stent; 2) a new technique for traversing tight pulmonary arterial corners for stent delivery using the "sheath-within-sheath" method; and 3) a new technique for recannulation and stent implantation in unilateral femoral venous occlusion.

Coronary artery fistula. Management and intermediate-term outcome after transcatheter coil occlusion.

During the last 2 decades, transcatheter occlusion of coronary artery fistulae has developed into a safe and effective therapy for children. This procedure avoids the need for open surgical repair and the attendant complications of cardiopulmonary bypass and median sternotomy. The long-term outcome in patients after transcatheter occlusion remains unknown. We describe the intermediate-term progress of 4 such patients after coil occlusion of coronary artery fistulae. Persistent coronary artery dilatation was present in all patients reviewed, as late as 4 years after occlusion.

Reconstruction of stenotic or occluded iliofemoral veins and inferior vena cava using intravascular stents: re-establishing access for future cardiac catheterization and cardiac surgery.

OBJECTIVES: The study evaluated the safety and efficacy of stent reconstruction of stenotic/occluded iliofemoral veins (IFV) and inferior vena cava (IVC). BACKGROUND: Patients with congenital heart defects and stenotic or occluded IFV/IVC may encounter femoral venous access problems during future cardiac surgeries or catheterizations. METHODS: Twenty-four patients (median age 4.9 years) underwent implantation of 85 stents in 22 IFV and 6 IVC. Fifteen vessels were severely stenotic and 13 were completely occluded. Although guide wires were easily passed across the stenotic vessels, occluded vessels required puncture through the thrombosed sites using a stiff wire or transseptal needle. Once traversed, the occluded site was dilated serially prior to stent implantation. RESULTS: Following stent placement, the mean vessel diameter increased from 0.9 +/- 1.6 to 7.4 +/- 2.6 mm (p < 0.05). Twenty-one of 28 vessels had long segment stenosis/occlusion requiring two to seven overlapping stents. Repeat catheterizations were performed in seven patients (9 stented vessels) at mean follow-up of 1.6 years. Seven vessels remained patent with mean diameter of 6.4 +/- 2.0 mm. Two vessels were occluded, but they were easily recanalized and redilated. Echocardiographic follow-up in two patients with IVC stents demonstrated wide patency. In four additional patients, a stented vessel was utilized for vascular access during subsequent cardiac surgery (n = 3) and endomyocardial biopsy (n = 1). Therefore, 13 of 15 stented vessels (87%) remained patent at follow-up thus far. CONCLUSIONS: Stenotic/obstructed IFV and IVC may be reconstructed using stents to re-establish venous access to the heart for future cardiac catheterization and/or surgeries.

Steerable control of the eustachian valve during transcatheter closure of secundum atrial septal defects.

Over the past decade there has been increased use of transcatheter devices for closure of secundum atrial septal defects. The presence of a large eustachian valve complicating transcatheter closure has not been described. We describe four patients with prominent eustachian valves, in three of whom we employed a simple technique to obtain control of the eustachian valve during device placement using transesophageal echo guidance.

Transcatheter closure of surgical shunts in patients with congenital heart disease.

The aim of this study was to review contemporary techniques, devices, and results of transcatheter occlusion of surgical shunts in 2 pediatric cardiac programs. Closure of superfluous surgical shunts may reduce cardiac work and risk of endocarditis. Previous studies have shown that transcatheter closure of shunts is feasible, but have not demonstrated acceptable efficacy or safety. In addition, the performance of new techniques and devices has not been reviewed. Between 1993 and 1998, 18 patients with congenital heart disease underwent transcatheter closure of 19 Blalock-Taussig shunts. Detachable and standard Gianturco coils and Gianturco-Grifka vascular occlusion devices were employed. All 19 shunts had complete closure. Eight shunts had initial placement of detachable coils. Five shunts had stents placed that bridged the pulmonary end of the shunts. These 5 and 4 additional shunts had closure by standard coils. Two shunts were closed with Gianturco-Grifka devices. There were no complications, no embolizations, and no requirement for surgery precipitated by the procedures. This review of contemporary techniques, devices, and results suggests that transcatheter occlusion of surgical shunts is effective and safe.