Efficient expression of stabilized mRNA PEG-peptide polyplexes in liver.

The expression efficiency in liver following hydrodynamic delivery of in vitro transcribed mRNA was improved 2000-fold using a codon-optimized mRNA luciferase construct with flanking 3' and 5' human ß-globin untranslated regions (UTR mRNA) over an unoptimized mRNA without ß-globin UTRs. Nanoparticle UTR mRNA polyplexes were formed using a novel polyacridine polyethylene glycol (PEG) peptide, resulting in an additional 15-fold increase in expression efficiency in the liver. The combined increase in expression for UTR mRNA PEG-peptide polyplexes was 3500-fold over mRNA lacking UTRs and PEG-peptide. The expression efficiency of UTR mRNA polyplex was 10-fold greater than the expression from an equivalent 1 µg dose of pGL3. Maximal expression was maintained from 4 to 24 h. Serum incubation established the unique ability of the polyacridine PEG-peptide to protect UTR mRNA polyplexes from RNase metabolism by binding to double-stranded regions. UTR mRNA PEG-peptide polyplexes are efficient nonviral vectors that circumvent the need for a nuclear uptake, representing an advancement toward the development of a targeted gene delivery system to transfect liver hepatocytes.

The uptake mechanism of PEGylated DNA polyplexes by the liver influences gene expression.

Two uptake mechanisms were identified for PEGylated DNA polyplex biodistribution to the liver. At a low polyplex dose, a rapid-uptake mechanism dominates, resulting in 60% capture by liver in 5 min, due to a saturable receptor-mediated process. Rapid-uptake led to the fast metabolism of polyplexes by liver (t1/2 = 2.1 h), correlating with a 1-µg pGL3 polyplex dose losing full transfection competency after 4 h in the liver. Dose escalation of either polyplex or poly(ethylene glycol) (PEG) peptide led to the saturation of rapid-uptake and revealed a delayed-uptake mechanism for polyplexes by liver. Delayed-uptake was characterized by the slower liver accumulation of 40% of the polyplex dose over 40 min, followed by slow metabolism (t1/2 = 15 h) and an extended time (12 h) for a 1-µg pGL3 polyplex dose, remaining fully transfection competent in the liver. The delayed-uptake mechanism is consistent with polyplexes crossing liver fenestrated endothelial cells to reach steady state in the space of Disse. The results describe how to control polyplex biodistribution to liver to avoid rapid-uptake and metabolism, in favor of delayed-uptake, to preserve polyplex transfection competency in the liver for up to 12 h.

Delayed gallium-67 uptake in renal atheroembolic disease.

The differentiation between atheroembolic disease (AED) and allergic interstitial nephritis (AIN) may pose a clinical challenge. Gallium scans have been proposed to identify AIN with good discriminating ability. We report herein a case of atheroembolic disease presenting as acute renal failure with persistent delayed uptake of gallium-67 by nuclear imaging. The distinction between AED and AIN could be made only with a renal biopsy, which confirmed the correct diagnosis. This case report and the presented review of the literature suggest that gallium scans are nonspecific and should not supplant renal biopsy for definitive histological diagnosis.

Percutaneous transluminal angioplasty of internal mammary arteries in patients with rest angina.

Angioplasty of the internal mammary artery (IMA) bypass graft has been shown to be a safe and effective revascularization procedure. However, angiographic and long term clinical outcomes in the high-risk group of patients presenting with rest angina has not been well documented. We report the results of IMA angioplasty in 20 patients with rest angina out of 614 (3.2%) who received a left IMA graft at our institution between April 1987 and September 1994. All patients were admitted with rest angina, 12 patients demonstrated persistent ischemia despite medical therapy, two patients were in heart failure, and one patient was in cardiogenic shock. Balloon angioplasty was successful in 15 of 20 patients (75%). Failed angioplasty was associated with either severe IMA tortuousity (three patients) or inability to cross the anastomotic stenosis with the guide wire (two patients). Each of these five patients required angioplasty of either the native left anterior descending artery or other saphenous vein grafts for clinical stabilization. No patient suffered a major complication (myocardial infarction, emergent coronary bypass surgery, death). Clinical follow-up was obtained in all 20 patients (6 months, 7 years, mean 27 months). Twelve patients (60%) were asymptomatic or had stable angina at follow-up, and 8 returned with anginal symptoms. Four patients required repeat angioplasty for disease in other vessels, two were treated medically for angina, one underwent repeat CABG, and cardiac transplantation was performed in one patient for refractory heart failure. Angiographic follow up was obtained in 10/15 (66%) successful angioplasty patients, and only one patient demonstrated restenosis at the treated site (10%). During follow up one patient developed an IMA stenosis at a previous dissection site in the body of the graft that was treated with angioplasty. These results suggest that IMA angioplasty in patients with rest angina is associated with excellent long term patency and clinical efficacy, as well as low procedural risk.

Percutaneous transluminal coronary angioplasty of the left main coronary artery in patients with rest angina.

Left main coronary angioplasty may be a therapeutic revascularization procedure for a subset of patients with symptomatic coronary artery disease. The purpose of this study is to report procedural outcomes and long-term clinical follow-up of 15 patients who underwent either protected or unprotected left main angioplasty for rest angina. These patients represent a cohort of unstable angina patients who were considered high risk for coronary artery bypass surgery. Ten of 15 patients had Canadian Heart Class IV angina, and three patients were hemodynamically unstable. Balloon angioplasty was successful in 4 patients, and one patient was treated with directional atherectomy. Initial angiographic success was achieved in 14 of 15 patients (93%). Major complications (myocardial infarction, emergent coronary artery bypass graft, death) occurred in one patient (6%); 73% of the patients were asymptomatic or had stable exertional angina at 6 months follow-up. One year survival was 87% (13 of 15). During the follow-up period six patients had repeat catheterization for recurrent angina. Four of these patients had left main restenosis and underwent successful repeat left main angioplasty. No patient had coronary bypass surgery during follow-up. This report suggests that left main angioplasty can be a safe and effective revascularization procedure for critically ill patients with unstable angina who are at high risk for coronary bypass surgery.

Platelet-induced vascular smooth muscle cell proliferation is modulated by the growth amplification factors serotonin and adenosine diphosphate.

BACKGROUND: Platelet-mediated mechanisms have been implicated in intimal lesion formation following vascular injury. Although the participation of peptide growth factors has been suspected in this process, little has been known about the possible mitogenic role of other platelet factors that are released at sites of vascular injury. METHODS AND RESULTS: We tested the hypothesis that platelet products, which are not peptide growth factors, are important modulators of the platelet-induced smooth muscle cell (SMC) proliferative response by acting as growth amplification factors. In these studies, cell proliferation was assessed by [3H]thymidine incorporation, flow cytometry, and direct cell counting. We examined the potential mitogenicity of several platelet products, including serotonin, ADP, norepinephrine, histamine, platelet-activating factor, the thromboxane A2 mimetic U46619, and bradykinin. Of the platelet products tested, serotonin and ADP induced a synergistic response with peptide growth factors. This synergy was greatest at low growth-factor concentrations. Addition of nonaggregated platelets to quiescent SMC cultures strongly stimulated cell proliferation. Since the addition of suramin to platelet-treated cultures markedly inhibited SMC proliferation, peptide growth factors are most likely the primary mitogens mediating this response. However, platelet-induced proliferation was also markedly reduced by the serotonin antagonists ketanserin and LY53857 (44%), and by the ADP antagonist apyrase (35%). CONCLUSIONS: Therefore, serotonin and ADP contribute significantly, in synergy with peptide growth factors, to the platelet-induced SMC proliferative response. We propose that in vivo serotonin and ADP act as amplification factors for SMC proliferation at sites of vascular injury.

Percutaneous transluminal angioplasty of saphenous vein grafts for medically refractory unstable angina.

OBJECTIVES: We attempted to answer the question, Is balloon angioplasty a reasonable alternative to repeat coronary artery bypass graft surgery in patients with previous coronary bypass graft surgery, medically refractory unstable angina and vein graft lesions? BACKGROUND: Patients with medically refractory unstable angina need revascularization. Patients with previous coronary artery bypass graft surgery and medically refractory angina are at "high risk" for adverse outcomes with repeat coronary bypass graft surgery. Conversely, patients with angioplasty of old vein grafts are also at "high risk" for adverse outcomes. METHODS: Balloon angioplasty of 89 lesions in saphenous vein grafts was performed in 75 consecutive patients with medically refractory unstable angina. Of these 75 patients, 24 (32%) had myocardial infarct within 30 days, 23 (31%) had left ventricular ejection fraction < 0.35, and 50 (67%) had major comorbidity. Patients underwent standard balloon angioplasty with aggressive use of intravenous and intracoronary heparin, urokinase, nitroglycerin, oral aspirin, calcium channel blocking agents and coumadin. RESULTS: Angiographic success (reduction of stenosis < or = 50% without major complication) was seen in 84 of 89 lesions. Clinical success (angiographic success plus hospital discharge without major complication) was seen in 70 of 75 patients. During index hospitalization, two patients (3%) died, two (3%) had nonfatal infarcts, and one (1%) had emergency reoperation (coronary bypass graft surgery). In late follow up (3 to 66 months), 14 (20%) patients were lost to follow-up, 17 (23%) had repeat percutaneous transluminal coronary angioplasty, 2 (3%) had late bypass graft reoperation, 18 (25%) had late death, and 1 (< 1%) had a heart transplant. Of the 41 patients alive after one or more angioplasties, 25 have little or no angina, and 16 have occasional or more angina. We compared long-term survival rate in these 75 patients with a cohort of patients with high risk, unstable angina from the Veterans Affairs Surgical Registry (2,570 patients). The 30-day survival rate was better in patients with coronary angioplasty (97% vs. 92%, p < 0.05), but by 6 months there was no difference, and by 5 years a trend toward a higher survival rate with coronary artery bypass graft surgery was seen. CONCLUSIONS: Balloon angioplasty of saphenous vein grafts with aggressive adjunctive pharmacotherapy is a reasonable alternative to repeat coronary bypass graft surgery in patients with medically refractory unstable angina, previous coronary bypass graft surgery and saphenous vein narrowing.