Predictors of Occlusion of Hepatic Blood Vessels after Irreversible Electroporation of Liver Tumors

PurposeTo examine predictors of midterm occlusion in portal and hepatic veins within or adjacent to the ablation zone after irreversible electroporation (IRE) of liver tumors.Materials and MethodsThis retrospective cohort analysis included 39 patients who underwent CT-guided IRE of liver tumors. Vessels within or adjacent to the ablation zone were identified on CT images acquired immediately after the procedure, and the positional relationships with the ablation zone (within/adjacent), locations (proximal/distal), and diameters (< 4 mm or ≥ 4 mm) were evaluated. Using contrast-enhanced follow-up scans, each vessel was classified as patent, stenosed, or occluded. Associations between vessel occlusion and each variable were investigated.ResultsOverall, 33 portal veins and 64 hepatic veins were analyzed. Follow-up scans showed occlusion in 12/33 (36.7%) portal veins and 17/64 (26.6%) hepatic veins. Vessels within the ablation zone were occluded significantly more frequently than vessels adjacent to the ablation zone (portal: 55.6% [10/18] vs 13.3% [2/15], P = .04; hepatic: 45.4% [15/33] vs 6.4% [2/31], P = .011). Vessels with a diameter < 4 mm were also occluded significantly more frequently than vessels with a diameter ≥ 4 mm (portal: 72.7% [8/11] vs 18.1% [4/22], P = .011; hepatic: 54.8% [17/31] vs 0% [0/33], P < .001). The respective positive and negative predictive values for occlusion of vessels categorized as both within and < 4 mm were 88% (7/8) and 82% (20/25) for portal veins and 79% (15/19) and 96% (43/45) for hepatic veins.ConclusionsMidterm vessel occlusion after liver IRE could be predicted with relatively high accuracy by assessing ablation location and vessel diameter.     

High-Frequency Irreversible Electroporation Using 5,000-V Waveforms to Create Reproducible 2- and 4-cm Ablation Zones—A Laboratory Investigation Using Mechanically Perfused Liver

PurposeTo investigate if high-frequency irreversible electroporation (H-FIRE) treatments can be delivered at higher voltages and with greater energy delivery rates than currently implemented in clinical irreversible electroporation protocols.Materials and MethodsTreatments using 3,000 V and 5,000 V were administered to mechanically perfused ex vivo porcine liver via a single applicator and grounding pad (A+GP) as well as a 4-applicator array (4AA). Integrated energized times (IET) 0.01–0.08 seconds and energy delivery rates 25–300 μs/s were investigated. Organs were preserved at 4°C for 10–15 hours before sectioning and gross analysis using a metabolic stain to identify the size and shape of ablation zones.ResultsA+GP ablations measured between 1.6 cm and 2.2 cm, which did not increase when IET was increased from 0.02 seconds to 0.08 seconds (P > .055; range, 1.9–2.1 cm). Changes in tissue color and texture consistent with thermal damage were observed for treatments with energy delivery rates 50–300 μs/s, but not for treatments delivered at 25 μs/s. Use of the 4AA with a 3-cm applicator spacing resulted in ablations measuring 4.4–4.9 cm with energy delivery times of 7–80 minutes.ConclusionsH-FIRE treatments can rapidly and reproducibly create 2-cm ablations using an A+GP configuration. Treatments without thermal injury were produced at the expense of extended treatment times. More rapid treatments resulted in ablations with varying degrees of thermal injury within the H-FIRE ablation zone. Production of 4-cm ablations is possible using a 4AA.     

Synergistic effect of low dose Cyclosporine A and human interleukin 10 overexpression on acute rejection in rat lung allotransplantation

Objective: Electroporation mediated transfer of plasmid DNA into peripheral muscle results in high transfection efficiency. The aim of this study was to investigate the effect of gene transfer of human IL-10 (hIL-10) into the tibialis anterior muscle (MTA) in combination with low dose Cyclosporine A (CsA) on acute rejection of lung allografts in the rat. Methods: Lung allotransplantation was performed from male BN donor to male Fisher F344 rats. Gene transfer was achieved by intramuscular injection into the MTA of the recipient followed by electroporation (4×20 ms impulses at 200 V/cm) 24 h prior to the transplantation. Group A (n=5) received CsA (2.5 mg/kg bw ip) for 5 days post-transplant and group B (n=5) 2.5 μg of PCIK hIL-10 (plasmid expression vector containing human CMV immediate early gene promoter and enhancer) and a low dose CsA (2.5 mg/kg bw i.p.). Graft function was assessed by blood gas at day 5 after exclusion of the native lung. Animals were sacrificed and blood was drawn to measure serum hIL-10 levels (ELISA) and tissue was sampled for histological grading of rejection. Results: Local expression of hIL-10 was confirmed at the mRNA level by in situ hybridization. All group A control animals showed severe signs of rejection. At day 5 all grafts in group B showed good gas exchange mean PaO2 233±123 mmHg, vs 44±8 mmHg in group A. Histological examination revealed moderate to severe rejection in all animals in group A (IIIB, ISHLT) in contrast to low moderate rejection in group B (II–IIIA). hIL-10 serum levels on day 5 were 14±7 pg/ml in group B vs. 0 in group A. Conclusions: Electroporation mediated hIL-10 overexpression in a peripheral muscle of the recipient in combination with low dose CsA reduces acute rejection in this model of rat lung allotransplantation.     

S波段高功率微波辐射对原代培养乳鼠心肌细胞的影响

目的研究S波段高功率微波（HPM）辐射后乳鼠心肌细胞的损伤效应及其机制。方法采用平均功率密度为5～30mW/cm^2的S波段HPM模拟源辐射原代培养的乳鼠心肌细胞，应用流式细胞术、原子力显微镜观察辐射后心肌细胞凋亡和坏死率、[Ca^2＋]。及细胞膜形态。结果10—30mW／cm^2的S波段HPM辐射后6h，心肌细胞凋亡和坏死率增加（P〈0．05或P〈0．01），且随辐射剂量增大，坏死细胞增多；30mW/cm^2 HPM辐射后即刻[Ca^2＋]i升高（P〈0．01），细胞膜发生穿孔。结论10—30mW/cm^2的S波段HPM辐射可造成心肌细胞凋亡和坏死，[Ca^2＋]；升高以及细胞膜穿孔，是S波段HPM辐射致心肌细胞损伤的重要机制。     

Ethanol-Induced Enhancement of Cocaine Bioactivation and Irreversible Protein Binding: Evidence Against a Role of Cytochrome P-450IIE1

Chronic ethanol consumption potentiates cocaine-induced liver injury in rodents. Since cocaine has to be bioactivated by a cytochrome P-450-dependent N-oxidative pathway to exert its hepatotoxic effects, we studied the role of the ethanol-inducible P-450IIE1 for cocaine metabolism. Male Sprague-Dawley rats were pretreated with either a liquid diet containing ethanol (30% of calories) for 4 weeks or injected with pyrazole (200 mg/kg/day, ip, for 3 days). Both agents induced microsomal p-nitrophenol hydroxylation which is a probe for the catalytic activity of P-450IIE1. However, only ethanol, but not pyrazole, increased both microsomal cocaine N-demethylase activity (by 47%) and the extent of irreversible binding of [3H]-cocaine to microsomal proteins (by 100%), which was taken as a quantitative endpoint for the formation of a reactive metabolite. Cocaine N-demethylation and irreversible protein binding of cocaine were not inhibited by P-450IIE1 isozyme-selective substrates, nor was the rate of cocaine metabolism and binding decreased by functionally active polyclonal anti-rat P-450IIE1 antibodies. Furthermore, pyrazole pretreatment sensitized cultured hepatocytes to the glutathione-dependent cytotoxic effects of nontoxic concentrations of cocaine. These results indicate that (a) cocaine is not a major substrate for the ethanol-inducible P-450IIE1, (b) the enhancing effects of ethanol on cocaine bioactivation may be due to induction of other P-450 isoforms, and (c) induction of P-450IIE1 may potentiate cocaine-induced hepatocellular toxicity in vitro independently of cocaine metabolism, e.g., by P-450IIE1-dependent oxidative stress.     

陡脉冲电场对荷瘤BALB/c小鼠癌细胞杀伤效应的实验研究

观测陡脉冲电场对荷瘤BALB/c小鼠癌细胞的杀伤效应。处理后的癌细胞在光镜下表现为细胞核固缩、核碎裂、核溶解;在电镜下表现为异染色质增加,边集,呈团块样改变,细胞质肿胀,脂滴数量增加,质膜破裂;核固缩,碎裂,线粒体肿胀。实验揭示了陡脉冲电场能有效地杀伤癌细胞,明显抑制了荷瘤小鼠恶性肿瘤的生长、增殖,有着良好的应用前景。     

电穿孔增强核酸疫苗免疫反应的研究进展

核酸疫苗自诞生以来在肿瘤、感染性疾病和自身免疫性疾病等方面显示出其独特的优越性。目前,研究如何增强核酸疫苗的免疫原性是一大热点,在众多方法中,电穿孔通过增加细胞对DNA的摄取,使表达的目的抗原增多,从而明显地增强了核酸疫苗诱导的免疫反应。就电穿孔技术在核酸疫苗研究领域的应用作一综述。     

Effects of the irreversible α-adrenoceptor antagonists phenoxybenzamine and benextramine on the effectiveness of nifedipine in inhibiting α 1- and α 2-adrenoceptor mediated vasoconstriction in pithed rats

Summary In pithed normotensive rats, i.v. injection of the selective ₁-adrenoceptor agonist cirazolien produced vasoconstriction which was largely resistant to inhibition by nifedipine. On the other hand, the pressor effects of the selective ₁-adrenoceptor agonists St 587 and Sgd 101/75 were much more effectively blocked by nifedipine, although not as effectively as the pressor effects to the selective ₂-adrenoceptor agonist B-HT 920. The sensitivity to inhibition of vasoconstriction in pithed rats to the different agonists increased in the order cirazoline St 587 ₁-, but not to ₂-adrenoceptor activation was dose-dependently enhanced. The potency of nifedipine to inhibit ₁-vasoconstriction by cirazoline, St 587 and Sgd 101/75 was increased maximally to the level of efficacy at which nifedipine antagonized B-HT 920-induced vasoconstriction. The dose of phenoxybenzamine required to maximally increase the potency and efficacy of nifedipine to antagonize vasoconstriction of the ₁-adrenoceptor agonists was inversely related to the level of sensitivity to blockade by nifedipine of the vasoconstriction they produced. In contrast, pretreatment of rats with the irreversible antagonist, benextramine (10 mg/kg, i.v., –100 to –60 min) did not increase the potency or efficacy of nifedipine to antagonize vasoconstriction to cirazoline, St 587, Sgd 101/75 or B-HT 920, despite irreversible blockade of ₁- and ₂-adrenoceptors. These data suggest that phenoxybenzamine, but not benextramine, selectively inhibits the ₁-adrenoceptor mediated vasoconstrictor mechanism that is independent of influx of extracellular calcium. Moreover, the results show that the existence of receptor reserve or the number of ₁-adrenoceptors activated does not determine the relative contribution of calcium influx-independent mechanisms in ₁-adrenoceptor-mediated vasoconstriction.Preliminary data were communicated at the Joint Meeting of the French and German Pharmacological and Toxicological Societies, Freiburg i. Br., September 19–22, 1983 (Timmermans et al. 1983a) and at the Winter Meeting of the British Pharmacological Society, London, January 1984 (De Jonge et al. 1984)     

Transdermal Delivery of Metoprolol by Electroporation

Electroporation, i.e., the creation of transient pores in lipid membranes leading to increased permeability, could be used to promote transdermal drug delivery. We have evaluated metoprolol permeation through full thickness hairless rat skin in vitro following electroporation with an exponentially decaying pulse. Application of electric pulses increased metoprolol permeation as compared to diffusion through untreated skin. Raising the number of twin pulses (300 V, 3 ms; followed after 1 s by 100 V, 620 ms) from 1 to 20 increased drug transport. Single pulse (100 V, 620 ms) was as effective as twin pulse application (2200 V, 1100 V or 300 V, 3 ms; followed after 1 s by 100 V, 620 ms). In order to investigate the effect of pulse voltage on metoprolol permeation, 5 single pulses (each separated by 1 min) were applied at varying voltages from 24 to 450 V (pulse time 620 ms). A linear correlation between pulse voltage and cumulative metoprolol transported after 4 h suggested that voltage controls the quantity of drug delivered. Then, the effect of pulse time on metoprolol permeation was studied by varying pulse duration of 5 single 100 V pulses from 80 to 710 ms (each pulse also separated by 1 min). Cumulative metoprolol transported after 4 h increased linearly with the pulse time. Therefore, pulse time was also a control factor of the quantity of drug delivered but to a lesser extent than the voltage at least at 100 V. The mechanisms behind improved transdermal drug delivery by electroporation involved reversible increased skin permeability, electrophoretic movement of drug into the skin during pulse application, and drug release from the skin reservoir formed by electroporation. Thus, electroporation did occur as shown by the increased transdermal permeation, on indicator of structural skin changes and their reversibility. Electroporation has potential for enhancing transdermal drug delivery.