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K L March

Publications and source records attributed to K L March.

At least 19 recordsLinked to original sources
PubMed2001

Liquid-filled balloon brachytherapy using (68)Ga is effective and safe because of the short 68-minute half-life: results of a feasibility study in the porcine coronary overstretch model.

BACKGROUND: Liquid-filled balloons for coronary brachytherapy provide significant advantages over solid sources in dose homogeneity but carry the risk of life-threatening radiointoxication after balloon rupture and laboratory contamination in case of a spill. We hypothesized that the positron emitter (68)Ga, with a half-life of only 68 minutes, was well suited to overcome these safety obstacles while providing full therapeutic efficacy. METHODS AND RESULTS: The feasibility, efficacy, and safety of (68)Ga liquid-filled balloon brachytherapy were investigated in the porcine coronary overstretch model. Four groups of 5 balloon-induced coronary lesions were irradiated with 8, 12, 16, and 24 Gy targeted to the adventitia. Ten unirradiated lesions served as controls. Segments treated with 16 or 24 Gy exhibited marked suppression of neointimal proliferation at 28-day follow-up, with quantitative parameters of intraluminal proliferation reduced to <20%. This beneficial effect was not compromised by untoward edge effects. Uninjured but irradiated vessels did not show histological signs of radiation damage. The (68)Ga whole-body dose due to balloon rupture was estimated to be 5 rem/50 mCi treatment activity and compared favorably with that of (188)Re (78 rem/50 mCi). CONCLUSIONS: (68)Ga positron radiation suppresses neointimal proliferation at doses of 16 and 24 Gy. This biological efficacy, coupled with the attractive safety profile, suggests the selection of (68)Ga as an attractive isotope for liquid-filled balloon brachytherapy.

Animals↗
PubMed2001

Improved laser-assisted vascular tissue fusion using light-activated surgical adhesive in a porcine model.

Newly developed light-activated surgical adhesives have been investigated as a substitute to traditional protein solders for vascular tissue fusion without the need for sutures. Porcine carotid arteries (n = 6) and femoral arteries (n = 6) were exposed, and an incision was made in the arterial walls using a 16G needle. The surgical adhesive, composed of a poly(L-lactic-co-glycolic acid) scaffold doped with the traditional protein solder mix of porcine serum albumin and indocyanine green dye, was used to close the incisions in conjunction with an 805 nm diode laser. Blood flow was restored to the vessels immediately after the procedure and the incision sites were checked for patency. The new adhesives were flexible enough to be wrapped around the vessels while their solid nature avoided the problems associated with "runaway" of the less viscous liquid protein solders widely used by researchers. The strength and hemostatic abilities of the new surgical adhesives were evaluated in the context of arterial pressure, persistence of hemostasis and presence of any inflammatory reaction after 3 days. After this evaluation period, the surgical procedure was repeated on the carotid arteries (n = 6) and femoral arteries (n = 6) of three additional animals that had been heparinized prior to surgery to closer approximate the conditions seen in a typical vascular surgical setting. The patency rate of both the unheparinized and heparinized vessels was 100% at 3 days post-operative with evidence of intraluminal thrombosis seen in only one of the repaired vessels. The adhesive technique also compared favorably with a previous study conducted using conventional suture techniques. Repairs formed with the adhesive technique were achieved more rapidly than suturing, and acute leakage was observed less frequently. Finally, the adhesive technique produced better histology than the suture technique, suggesting that it has great promise as an alternative to suturing. These initial results indicate that laser-assisted vascular repair using the new adhesives is safe, easy to perform, and contrary to conventional suturing, provides an immediate leak-free closure. In addition, the flexible and moldable nature of the new adhesives allows them to be tailored to a wide range of tissue geometries, thus greatly improving the clinical applicability of laser-assisted tissue repair.

Animals↗
PubMed2001

Advantages of short-lived positron-emitting radioisotopes for intracoronary radiation therapy with liquid-filled balloons to prevent restenosis.

UNLABELLED: Balloon catheters filled with liquid radioisotopes provide excellent dose homogeneity for intracoronary radiation therapy but are associated with risk for rupture or leakage. We hypothesized that the safety of liquid-filled balloons may be improved once positron emitters with half-lives below 2 h are used instead of the high-energy beta-emitters 166Ho, 186Re, or 188Re, all of which have a longer half-life of at least 17 h. METHODS: To support this concept, the suitability of 18F (half-life, 109.8 min), 68Ga (half-life, 67.6 min), 11C (half-life, 20.4 min), 13N (half-life, 9.97 min), and 15O (half-life, 2.04 min) for intracoronary radiation therapy was evaluated. Potential tissue penetration of positron radiation was assessed in a series of phantom experiments using Gafchromic film. Antiproliferative efficacy of positrons emitted by 68Ga was investigated in vitro using cultured bovine aortic smooth muscle cells (BASMCs), and was compared with gamma-radiation emitted by 137Cs. To characterize the remaining risk, we estimated radiotoxicity after accidental intravascular balloon rupture on the basis of tabulated isotope-specific doses (ICRP 53) and compared these values with 188Re. RESULTS: Half-dose depth of tissue penetration measured in phantom experiments was 0.29 mm for 18F, 0.42 mm for 11C, 0.54 mm for 13N, 0.79 mm for 15O, and 0.9 mm for 68Ga. Irradiation of cultured BASMCs with positron radiation (68Ga) induced dose-dependent inhibition of proliferation with complete proliferative arrest at doses exceeding 6 Gy. ED(50) and ED(80) were 2.5 +/- 0.4 Gy (mean +/- SD) and 4.4 +/- 0.8 Gy, respectively. Antiproliferative efficacy was equal to that of the 662-keV gamma-radiation emitted by 137Cs (ED(50), 3.8 +/- 0.2 Gy; ED(80), 8.0 +/- 0.3 Gy). Estimates made for patient whole-body and organ doses were generally below 50 mSv/1.85 GBq for all investigated positron emitters. The same dose estimates for 188Re were 6-20 fold higher. CONCLUSION: Among the studied radioisotopes, 68Ga is the most attractive source for liquid-filled balloons because of its convenient half-life, sufficient positron energy (2.92 MeV), documented antiproliferative efficacy, and uncomplicated availability from a radioisotope generator. The safety profile for 68Ga is significantly better than that of 188Re, which suggests this radioisotope should be evaluated further in preclinical studies.

Animals↗
PubMed2000

Intrapericardial paclitaxel delivery inhibits neointimal proliferation and promotes arterial enlargement after porcine coronary overstretch.

BACKGROUND: Catheter-based intrapericardial (IPC) delivery of therapeutic agents has recently been demonstrated. Paclitaxel is known to inhibit vascular smooth muscle cell proliferation. This study examined the effect of IPC instillation of paclitaxel on neointimal proliferation after balloon overstretch of porcine coronary arteries. METHODS AND RESULTS: Overstretch injury of coronary arteries was followed by IPC administration of micellar paclitaxel at low dose (LD, 10 mg; n=6) or high dose (HD, 50 mg; n=7) or of control micelles (50 mg, n=5). Animals were euthanized 28 days after balloon dilation. Arterial injury indices were no different among the groups. The neointimal area, maximal intimal thickness, and adventitial thickness were significantly reduced in both LD (0.47+/-0.04 mm(2), 0.43+/-0.03 mm, and 0.35+/-0.02 mm, respectively) and HD (0.51+/-0.06 mm(2), 0.42+/-0.03 mm, and 0. 38+/-0.03 mm, respectively) paclitaxel groups compared with the control group (0.79+/-0.07 mm(2), 0.56+/-0.02 mm, and 0.47+/-0.02 mm, respectively; P:<0.001). Meanwhile, the vessel circumference measured at the external elastic lamina of paclitaxel-treated vessels was significantly larger than the control circumference. Apoptotic cells were found in the neointima. The apoptotic cell percentage was not different between the control (1.72%) and LD (2. 31%) groups but was higher in the HD group (7.07%, P:<0.0001 versus control and LD groups). Immunostaining for matrix metalloproteinase-2 revealed concurrent reduction in the HD group compared with the control and LD groups. CONCLUSIONS: IPC space delivery of a single dose of paclitaxel significantly reduces vessel narrowing in this balloon-overstretch model. This effect is mediated by reduction of neointimal mass as well as positive vascular remodeling.

Angioplasty, Balloon, Coronary↗
PubMed2000

Differences in the effects of HMG-CoA reductase inhibitors on proliferation and viability of smooth muscle cells in culture.

We investigated the influence of lovastatin, simvastatin and pravastatin on proliferation and viability of vascular smooth muscle cells (SMC) in vitro and studied the effects of lovastatin on a mouse SMC line transgenic for a temperature-sensitive mutant of SV40 large T antigen (TAg), known to inhibit the function of p53 and pRb family members. We found that lovastatin and simvastatin inhibited cell proliferation by provoking G0/G1 phase arrest with concomitant depression of the proliferation antigen Ki-67/MIB-1. Lovastatin at high concentrations of 20 micromol/l caused cell death in the presence of serum but not under serum starved conditions, which was verified on the basis of increased DNA strand breaks, decreased DNA content and morphological alterations seen by electron microscopy. Cell death was also found for simvastatin, whereas pravastatin did not exhibit antiproliferative or cytotoxic effects. Mouse SMC transgenic for TAg did not show any impaired sensitivity to the antiproliferative and cell death inducing effect of lovastatin, but both effects could be antagonized by the supplementation of mevalonate. The data indicate that antiproliferative and cytotoxic effects of lovastatin are caused by the using up of products of mevalonate metabolism and do not require the presence of p53 or pRb.

Animals↗
PubMed2000

Studies of renal injury III: lipid-induced nephropathy in type II diabetes.

UNLABELLED: Studies of renal injury III: Lipid-induced nephropathy in type II diabetes. BACKGROUND: Nephrotoxicity from elevated circulating lipids occurs in experimental and clinical situations. We tested the hypothesis that lipid-induced nephropathy causes advanced renal failure in rats with type II diabetes and dyslipidemia. METHODS: First generation (F1) hybrid rats derived from the spontaneous hypertensive heart failure rat (SHHF/Gmi-fa) and the LA/NIH-corpulent rat (LA/N-fa) were studied for 41 weeks while being on specific diets. Group 1 (14 rats) ingested 11.5% protein, 47.9% fat, and 40.6% carbohydrate. Group 2 (8 rats) ingested 26.9% protein, 16.7% animal fat, and 56.4% carbohydrate, and group 3 (20 rats) ingested 20.2% protein, 40.4% soy and coconut oil, and 39.4% carbohydrate. RESULTS: Hyperglycemia was more severe in rat groups 1 and 2 than in group 3. In contrast, circulating cholesterol and hydroperoxide levels were highest in group 3, intermediate in group 2, and lowest in group 1. Group 3 had severe renal failure secondary to glomerulosclerosis and tubulointerstitial disease, with striking deposition of the lipid peroxidation stress biomarker 4-hydroxynonenal in glomeruli and renal microvessels. Moreover, in group 3, increased arterial wall thickness also connoted vascular injury. In contrast, the glycoxidation stress biomarkers pentosidine and carboxymethyl-lysine were preferentially localized to renal tubules of hyperglycemic rats in groups 1 and 2 and did not segregate with the most severe renal injury. Glomerular and interstitial fibrosis was accompanied by proportional increases in renal transforming growth factor-beta1 levels, which were threefold higher in the hypercholesterolemic rats of group 3 than in the hyperglycemic rats of group 1. CONCLUSIONS: Acquisition of non-nodular glomerular sclerosis and tubulointerstitial disease is dependent on lipoxidation stress in rats with type II diabetes. On the other hand, in the absence of hypercholesterolemia, prolonged glycoxidation stress does not appear to be uniquely nephrotoxic.

Animals↗
PubMed1999

Hyperplasia in multiple smooth muscle tissues in transgenic mice expressing a temperature-sensitive SV40 T-antigen under the control of smooth muscle alpha-actin regulatory sequences.

Control of smooth muscle cell (SMC) proliferation is of fundamental importance in the development and pathology of the vasculature. To derive vascular SMC with conditional inactivation of negative cell cycle regulatory proteins in the context of smooth muscle protein expression, a 3.4 kb fragment of the mouse SMC alpha-actin promoter was used to target a temperature-sensitive mutant SV40 T antigen (tsA58) to smooth muscle in transgenic mice. Mice with this genotype display a heritable phenotype of abnormal SMC proliferation in the central tail artery, vasa deferentia, seminal vesicles, prostate, and uterus, with the latter resembling uterine leiomyomatosis and prostatic hypertrophy. Neither the aorta nor other viscera manifested abnormal proliferation. Cultures from aorta, vas deferens, seminal vesicle, and kidney tissue were characterized with regard to protein expression, stability, and matrix remodelling capacity. The alpha-actin content/cell was up to 3-4-fold higher, as well as more stable than in primary SMC cultures, suggesting successful selection for propagation of cells expressing this differentiation marker. All cells displayed enhanced growth at the permissive temperature. As an initial functional assessment, the cells were compared to non-transformed mouse aortic SMC with respect to the ability to remodel collagen gel matrices, and demonstrated conservation of this physiologic function. This in vivo analysis of the SMC alpha-actin promoter supports a broader range of smooth muscle-directed expression activity than previously recognized, and establishes the feasibility of its use to direct transgene expression to vascular as well as genito-urinary smooth muscle. The targeted expression of the tsA58 T antigen has yielded transgenic animals with several manifestations of smooth muscle hyperplasia; these animals have in turn permitted the derivation of several murine SMC lines with phenotypic stability and conditionally-modulated proliferation. These cells will allow expansion of derivative transfected smooth muscle cell lines under permissive conditions, as well as oncogene inactivation at the restrictive temperature when desired for functional studies.

Actins↗
PubMed1999

Pharmacokinetics and consistency of pericardial delivery directed to coronary arteries: direct comparison with endoluminal delivery.

BACKGROUND AND HYPOTHESIS: Pharmacologic modulation of the contents of the pericardial space has been shown to influence the response of coronary arteries to balloon injury. Endoluminal (EL) local delivery of various drugs into coronaries has been found to be limited by short residence time, as well as by highly variable deposited agent concentration. We hypothesized that compounds placed into the pericardial space (P) would penetrate into coronary tissue with greater consistency than seen after EL delivery and provide for prolonged coronary exposure to agents. METHODS AND RESULTS: 125I-labeled basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF), albumin, or 131I-labeled diazeniumdiolated albumin (NONO-albumin) were delivered as model/therapeutic proteins into the porcine pericardial space (n = 15 pigs) or into coronaries using an EL delivery catheter (n = 48 arteries). In subjects receiving 125I-labeled proteins, the delivery target or mid-regions of the left anterior descending (LAD) and left circumflex (LCx) arteries were harvested at 1 h or 24 h for gamma-counting and autoradiography, and fractional intramural delivery (FID) or retention measured as percent agent in 100 mg artery/agent in infusate for both time points. In the animals receiving 131I-labeled NONO-albumin, serial gamma imaging was employed to evaluate the rate of redistribution in individual animals following either pericardial or endoluminal delivery. At 1 h, FID values ranged from 0.00064 to 0.0052% for P delivery (median 0.0022%), and from 0.00021 to 6.7 for EL delivery (median 0.27%). At 24 h, FID values ranged from 0.00011 to 0.003 for P delivery (median 0.0013), and from 0.0002 to 1.4 for EL delivery. The estimated T1/2 for bFGF redistribution from the vascular tissue was 22 h (P) and 7 h (EL), respectively, while the directly determined T1/2 values for NONO-albumin redistribution from the delivery region were 22.2 h (P) and 2.5 h (EL). CONCLUSIONS: These data show that pericardial fluid contents can access coronary arteries with intramural concentrations which typically vary by 10-15-fold, while EL delivery results in a remarkably wide intramural concentration range with up to 33,000-fold variability. The apparent redistribution rate is more rapid following EL delivery, possibly due to sustained diffusive tissue loading from the pericardial space. Pericardial delivery appears to offer substantial advantages over EL administration with respect to residence time and reproducibility.

Albumins↗
PubMed1999

Efficient in vivo catheter-based pericardial gene transfer mediated by adenoviral vectors.

Adenoviral vectors are promising agents for a number of in vivo gene therapy applications including diseases of the heart and coronary vessels. Efficient intravascular gene transfer to specific sites has been achieved in occluded vessels, but otherwise is hampered by the effect of blood flow on localized vector uptake in the vessel wall. An alternative delivery approach to coronary arteries is the expression of diffusible gene products into the pericardial space surrounding the heart and coronary arteries. However, in vivo pericardial access is comparatively difficult and has been limited to surgical approaches. We hypothesized that efficient adenovirus-mediated gene expression in pericardial lining mesothelium could be achieved by transmyocardial vector delivery to the pericardium. To evaluate this concept, a hollow, helical-tipped penetrating catheter was used to deliver vector-containing fluid directly into the intrapericardial space. The catheter was introduced percutaneously in anesthetized mongrel dogs, advanced into the right ventricle, and the tip passed through the apical right ventricular myocardium under direct radiographic visualization until the open end of the catheter tip resided in the intrapericardial space. Adenoviral vectors expressing either nuclear-localizing beta-galactosidase, cytoplasmic luciferase, or secreted human alpha 1AT reporters (Av1nBg, Av1Lu, or Av1Aa, respectively) were instilled through the catheter into the intrapericardial space. Three days later the animals were sacrificed and reporter gene expression was evaluated in pericardium, epicardium, and multiple other tissues. In animals receiving Av1nBg, beta-galactosidase activity was evident in most of the pericardial lining endothelium, up to 100% in many areas. In animals receiving Av1Lu, luciferase reporter activity was abundant in pericardial tissues, but near-background levels were observed in other organs. In animals receiving Av1Aa, human alpha 1AT was abundant (16-29 mg/ml) in pericardial fluid, but was undetectable in serum. All animals tolerated the procedure well with no electrocardiographic changes and no clinical sequelae. These observations demonstrate highly efficient adenovirus vector delivery and gene transfer and expression in the pericardium and support the feasibility of localized gene therapy via catheter-based pericardial approaches. We suggest that the pericardial sac may serve as a sustained-release protein delivery system for the generation of desired gene products or their metabolites for diffusion into the epicardial region.

Adenoviridae↗
PubMed1999

Establishment of a clinically correlated human pericardial fluid bank: evaluation of intrapericardial diagnostic potential.

The development of a clinically correlated human pericardial fluid bank and database is described. A unique feature of this registry is the availability of a large number of pericardial fluid samples for testing with respect to multiple factors and for correlation with angiographic findings and clinical syndromes expressed by the patients. The collection of data at the present time comprises frozen pericardial fluid samples obtained from patients who have undergone cardiac surgery; and historical, clinical, and laboratory data obtained from the patient records. Nearly 400 samples have been stored and analyzed thus far, with sample entry continuing. This registry is designed to evaluate the local factors that play a role in mediating or reflecting myocardial or coronary responses. Pathophysiologic processes of particular interest include restenosis, plaque ruptures, and angiogenesis. Study of the pericardial fluid bank should lead to enhanced understanding of molecular mechanisms, as well as to the explanation for the reasons underlying interpatient variability in these processes. It is further anticipated that this information might provide a foundation for the diagnostic use of pericardial fluid to individualize therapies targeting angiogenesis or plaque physiology.

Coronary Angiography↗
PubMed1999

NGF activates similar intracellular signaling pathways in vascular smooth muscle cells as PDGF-BB but elicits different biological responses.

The signaling pathways that regulate smooth muscle cell migration and proliferation are incompletely understood. Smooth muscle cells express at least 3 families of receptor tyrosine kinases that mediate cell migration: platelet-derived growth factor (PDGF) receptors, the trk family of neurotrophin receptors, and insulin-like growth factor 1 receptor. The neurotrophin, nerve growth factor (NGF), and insulin-like growth factor 1 induce the migration but not the proliferation of smooth muscle cells, whereas PDGF-BB stimulates both responses. To determine whether distinct signaling pathways downstream of receptor tyrosine kinases specifically mediate smooth muscle cell migration or proliferation, the ligand-induced activation of different signaling pathways in smooth muscle cells was examined. NGF induces prolonged activation of the Shc/MAP kinase pathway and phospholipase Cgamma compared with PDGF-BB. The activation of phosphatidylinositol-3 kinase, however, was 10-fold greater in response to PDGF-BB compared with NGF. Insulin-like growth factor 1 activates only phosphatidylinositol-3 kinase. Pharmacological inhibitors of phosphatidylinositol-3 kinase, Wortmannin and LY294002, inhibit PDGF-BB and NGF-induced migration, whereas an inhibitor of MAP kinase kinase, PD98059, has no effect. Our results suggest that (1) different receptor tyrosine kinases use similar patterns of activation of signaling pathways to mediate distinct biological outcomes of cell migration and proliferation, (2) NGF activates signaling proteins in smooth muscle cells similar to those activated during NGF-induced neuronal differentiation, and (3) the combinatorial effects of different signaling pathways are important for the regulation of smooth muscle cell migration and proliferation. Further studies using mutant trk receptors will help to define the signal transduction pathways mediating NGF-induced smooth muscle cell migration.

Animals↗
PubMed1998

Gene therapy for restenosis: getting nearer the heart of the matter.

Intensive work over the past decade has been directed to the study of vascular gene transfer as an approach to the unresolved problem of restenosis. This effort has resulted in a significant foundation of knowledge relative to the activities of potentially therapeutic gene products as well as the capabilities and limitations of vector systems and mechanical delivery modalities available for effecting the vascular expression of these gene products. In several instances, significant progress has been made by experiments highlighting unexpected difficulties and the need for more comprehensive understanding. It is thus now possible to clearly define and address specific challenges that must be overcome in order to make feasible progress from the preclinical to the clinical arena. The key challenges at present appear to include the evolution of clinically practical delivery methods that meet the kinetic requirements of achieving efficient gene transduction and the availability of vectors that maximize efficiency while minimizing undesirable host responses. Emerging data suggest that approaches to solving each of these issues may have recently been developed. Basic research evaluating these new delivery mechanisms and molecular vectors is essential to establish their true potential for use in the clinical arena.

Animals↗
PubMed1997

Activation of MAP kinase in vivo follows balloon overstretch injury of porcine coronary and carotid arteries.

Vascular restenosis involves contraction, proliferation, and remodeling of the arterial wall in response to overstretch injury. Mitogen-activated protein kinases (MAPKs) are implicated in both contraction and proliferation of vascular smooth muscle (VSM), and studies of porcine carotid arterial muscle strips have shown that mechanical stretch leads to the activation of the extracellular signal-regulated kinase (ERK) family of MAPKs in vivo. We, therefore, analyzed the acute effect of mechanical overstretch injury on ERK-MAPK (herein referred to simply as MAPK) activity in porcine coronary and carotid arteries in vivo. Balloon angioplasty catheters were inflated to 6 atm three times over 5 minutes at a balloon-artery ratio of 1.2:1 in either porcine coronary or carotid arteries. The arteries were snap-frozen after angioplasty, and MAPK activity was measured. Angioplasty of the left anterior descending (LAD, n = 5), left circumflex (LCx, n = 5), and carotid (n = 5) arteries effected an increase in MAPK activity compared with the activity in uninstrumented right coronary arteries (RCAs) or carotid arteries from the same animals used for controls. Balloon angioplasty of carotid arteries led to an increase in MAPK activity that was 7.7-fold over the activity in control arteries and comparable to the activity in stretched carotid arterial muscle strips in vivo. The increase in coronary artery kinase activity on angioplasty was variable from animal to animal. The increase in MAPK activity over that in control arteries ranged from 4.5- to 31.7-fold (mean +/- SEM, 10.7 +/- 5.3) in the LAD and 1.8- to 31.3-fold (mean +/- SEM, 9.7 +/- 5.7) in the LCx. There were no apparent inherent differences in the levels of MAPK activity in the three different types of coronary arteries (RCA, LAD, and LCx) without instrumentation. MAPK activation occurs rapidly during angioplasty, suggesting that this kinase may play an early role in initiating the injury response in both porcine coronary and carotid arteries. MAPKs may be key enzymes targeted to treat or prevent restenosis.

Angioplasty, Balloon↗
PubMed1996

Microparticle deposition in periarterial microvasculature and intramural dissections after porous balloon delivery into atherosclerotic vessels: quantitation and localization by confocal scanning laser microscopy.

Local delivery of pharmacologic or genetic agents with a porous balloon catheter offers a potential therapeutic approach to reducing restenosis and atherosclerosis and minimizing undesirable systemic toxicity. However, the delivery efficiency and intramural retention of liquid agents is low. The local intramural delivery and prolonged retention of 5 microns microparticles (MP) has been described previously. The current study was designed to evaluate the distribution of locally delivered MPs and to determine the effects of MP size and infusion pressure on intramural delivery efficiency. A 1% suspension of fluorescent, latex MPs (1 or 4.5 microns in diameter) was infused at either 3 or 6 atm into atherosclerotic rabbit femoral arteries (n = 32) immediately after angioplasty. Four groups of arteries were evaluated: 1) 1 micron MPs infused at 3 atm; 2) 1 micron MPs at 6 atm; 3) 4.5 microns MPs at 3 atm; and 4) 4.5 microns MP at 6 atm. The location of MPs was evaluated by fluorescent and light microscopy and confocal laser scanning microscopy. The tissue was dissolved and the delivered MPs quantified. All groups manifested numerous MPs within the vasa vasorum and periadventitial microvasculature, with a substantially lesser number within the neointimal and medial layers. The intramural deposition of the MPs was associated with dissection within the intima or media caused by the antecedent angioplasty or local delivery, indicating that an intact vessel wall is an anatomic barrier to MP delivery. The median values of fractional intramural delivery, defined as the percentage of infused MPs retained within the arterial wall, were 0.059%, 0.071%, 0.047%, and 0.062% for the groups 1 through 4, respectively (p not significant [NS]). The values of intramural particle concentration, expressed as the total number of MPs per weight of arterial tissue, were 55, 65, 1.5, and 1.2 x 10(4) MPs/mg for groups 1 through 4, respectively (p < 0.001 for 1 micron vs 4.5 microns MPs). Although more 1 microM MPs were delivered than 4.5 microns MPs, the fractional intramural delivery was unaffected by particle size or infusion pressure. The local delivery of MPs at atherosclerotic sites after angioplasty is characterized by fractional intramural delivery values similar to values of nonparticulate agents, with few MPs deposited into intima or media in the absence of a dissection caused by the antecedent angioplasty or the delivery procedure itself.

Angioplasty, Balloon, Coronary↗
PubMed1996

Vascular repair mechanisms after directional atherectomy or percutaneous transluminal coronary angioplasty in atherosclerotic rabbit iliac arteries.

Although directional atherectomy (DA) reduces the plaque burden, successful revascularization is not associated with a reduced restenosis rate when compared with percutaneous transluminal coronary angioplasty (PTCA). The purpose of this study was to compare and contrast the vascular response to DA-induced and PTCA-induced injury. Six to 8 weeks after induction of atherosclerosis, PTCA (n = 34) was performed in one iliac artery and DA in the other (n = 30). Arteries were obtained at 6 time points: 1, 3, 5, 7, 14, and 28 days. Eleven arteries that did not undergo an intervention acted as controls. Radiograms obtained before and after intervention and at euthanization were compared. Morphometric, histologic, and immunohistochemical analyses were performed. Both PTCA and DA resulted in an immediate increase in luminal diameter that subsequently decreased over the ensuing month. PTCA caused deep dissection (7 of 8 arteries), often extending to the adventitia, whereas stand alone DA resulted in deep cleft formation (4 of 5). Of the 30 arteries that underwent DA, 4 exhibited an increase in luminal diameter in the absence of tissue retrieval. Thrombus was observed in both the dissection planes and the clefts within the first 7 days, and cellular ingrowth was appreciated at 5 to 7 days. By 7 days the artery was repaired, and the histologic appearance of the arteries that had undergone PTCA could not be differentiated from the arteries that had undergone DA. Increased intimal and medial collagen and elastin was noted at 14 and 28 days. An increase in the area bordered by the external elastic lamina was observed in both groups. Although successful DA results in tissue removal and the production of a deep tissue cleft and PTCA causes a dissection, both produce a condition in which the arterial injury exposes the arterial media to blood, causing thrombus formation and inflammation with subsequent cellular accumulation into the thrombotic framework.

Aortic Dissection↗
PubMed1996

Evaluation of the concentration and bioactivity of adenovirus vectors for gene therapy.

Development of adenovirus vectors as potential therapeutic agents for multiple applications of in vivo human gene therapy has resulted in numerous preclinical and clinical studies. However, lack of standardization of the methods for quantifying the physical concentration and functionally active fraction of virions in these studies has often made comparison between various studies difficult or impossible. This study was therefore carried out to define the variables for quantification of the concentration of adenovirus vectors. The methods for evaluation of total virion concentration included electron microscopy and optical absorbance. The methods for evaluation of the concentration of functional virions included detection of gene transfer (transgene transfer and expression) and the plaque assay on 293 cells. Enumeration of total virion concentration by optical absorbance was found to be a precise procedure, but accuracy was dependent on physical disruption of the virion to eliminate artifacts from light scattering and also on a correct value for the extinction coefficient. Both biological assays for enumerating functional virions were highly dependent on the assay conditions and in particular the time of virion adsorption and adsorption volume. Under optimal conditions, the bioactivity of the vector, defined as the fraction of total virions which leads to detected target cell infection, was determined to be 0.10 in the plaque assay and 0.29 in the gene transfer assay. This difference is most likely due to the fact that detection by gene transfer requires only measurement of levels of transgene expression in the infected cell whereas plaque formation is dependent on a series of biological events of much greater complexity. These results show that the exact conditions for determination of infectious virion concentration and bioactivity of recombinant adenovirus vectors are critical and must be standardized for comparability. These observations may be very useful in comparison of data from different preclinical and clinical studies and may also have important implications for how adenovirus vectors can optimally be used in human gene therapy.

Adenoviridae↗
PubMed1996

Human surfactant protein A enhances attachment of Pneumocystis carinii to rat alveolar macrophages.

Pneumocystis carinii (PC) pneumonia remains one of the most important opportunistic pulmonary infections. The alveolar macrophage (AM) is likely the primary cell for recognition and removal of PC. The histopathology of PC pneumonia is characterized by a surfactant-like alveolar exudate. We hypothesize that surfactant protein A(SP-A), the major apoprotein of surfactant, mediates attachment of PC to rat AMs by acting as a ligand between the organism and the AM. In this study, attachment of PC was determined using (51)Cr-labeled PC incubated at 4 degrees C with normal rat AM monolayers in the presence or absence of human SP-A. SP-A significantly enhanced attachment of PC from 14.2 +/- 1.2% to 42.0 +/- 3.8% (P<0.05). This enhanced attachment was visualized and quantified morphologically with confocal microscopy. PC attachment by SP-A was calcium- and mannose-dependent as SP-A-mediated attachment was significantly reduced in the presence of EGTA and mannose to 13.1 +/- 1.6% and 19.3 +/- 2.6%, respectively (P<0.05). Addition of type V collagen and antibodies to SP-A also significantly reduced SP-A-mediated attachment to 4.9 +/- 1.2% and 10.1 +/- 1.2%, respectively (P<0.05). We conclude that SP-A can function as a ligand between PC and the AM and may represent an important detection and clearance mechanism of PC from the alveolar spaces.

Animals↗