Mechanisms and immunogenicity of nsPEF-induced cell death in B16F10 melanoma tumors.

Accumulating data indicates that some cancer treatments can restore anticancer immunosurveillance through the induction of tumor immunogenic cell death (ICD). Nanosecond pulsed electric fields (nsPEF) have been shown to efficiently ablate melanoma tumors. In this study we investigated the mechanisms and immunogenicity of nsPEF-induced cell death in B16F10 melanoma tumors. Our data show that in vitro nsPEF (20-200, 200-ns pulses, 7 kV/cm, 2 Hz) caused a rapid dose-dependent cell death which was not accompanied by caspase activation or PARP cleavage. The lack of nsPEF-induced apoptosis was confirmed in vivo in B16F10 tumors. NsPEF also failed to trigger ICD-linked responses such as necroptosis and autophagy. Our results point at necrosis as the primary mechanism of cell death induced by nsPEF in B16F10 cells. We finally compared the antitumor immunity in animals treated with nsPEF (750, 200-ns, 25 kV/cm, 2 Hz) with animals were tumors were surgically removed. Compared to the naïve group where all animals developed tumors, nsPEF and surgery protected 33% (6/18) and 28.6% (4/14) of the animals, respectively. Our data suggest that, under our experimental conditions, the local ablation by nsPEF restored but did not boost the natural antitumor immunity which stays dormant in the tumor-bearing host.

Vascular endothelial growth factor-A gene electrotransfer promotes angiogenesis in a porcine model of cardiac ischemia.

This study aimed to assess safety and therapeutic potential of gene electrotransfer (GET) as a method for delivery of plasmid encoding vascular endothelial growth factor A (VEGF-A) to ischemic myocardium in a porcine model. Myocardial ischemia was induced by surgically occluding the left anterior descending coronary artery in swine. GET following plasmid encoding VEGF-A injection was performed at four sites in the ischemic region. Control groups either received injections of the plasmid without electrotransfer or injections of the saline vehicle. Animals were monitored for 7 weeks and the hearts were evaluated for angiogenesis, myocardial infarct size and left ventricular contractility. Arteriograms suggest growth of new arteries as early as 2 weeks after treatment in electrotransfer animals. There is a significant reduction of infarct area and left ventricular contractility is improved in GET-treated group compared with controls. There was no significant difference in mortality of animals treated with GET of plasmid encoding VEGF-A from the control groups. Gene delivery of plasmid encoding VEGF-A to ischemic myocardium in a porcine model can be accomplished safely with potential for myocardial repair and regeneration.

Gene electro transfer of plasmid encoding vascular endothelial growth factor for enhanced expression and perfusion in the ischemic swine heart.

Myocardial ischemia can damage heart muscle and reduce the heart's pumping efficiency. This study used an ischemic swine heart model to investigate the potential for gene electro transfer of a plasmid encoding vascular endothelial growth factor for improving perfusion and, thus, for reducing cardiomyopathy following acute coronary syndrome. Plasmid expression was significantly greater in gene electro transfer treated tissue compared to injection of plasmid encoding vascular endothelial growth factor alone. Higher gene expression was also seen in ischemic versus non-ischemic groups with parameters 20 Volts (p<0.03), 40 Volts (p<0.05), and 90 Volts (p<0.05), but not with 60 Volts (p<0.09) while maintaining a pulse width of 20 milliseconds. The group with gene electro transfer of plasmid encoding vascular endothelial growth factor had increased perfusion in the area at risk compared to control groups. Troponin and creatine kinase increased across all groups, suggesting equivalent ischemia in all groups prior to treatment. Echocardiography was used to assess ejection fraction, cardiac output, stroke volume, left ventricular end diastolic volume, and left ventricular end systolic volume. No statistically significant differences in these parameters were detected during a 2-week time period. However, directional trends of these variables were interesting and offer valuable information about the feasibility of gene electro transfer of vascular endothelial growth factor in the ischemic heart. The results demonstrate that gene electro transfer can be applied safely and can increase perfusion in an ischemic area. Additional study is needed to evaluate potential efficacy.

Minimizing trauma to the upper airway: a ferret model of neonatal intubation.

Our objective was to determine whether an adult ferret can be intubated as many as 10 times per training session without resulting in trauma to the upper airway. In this program, 8 male ferrets rotated through intubation laboratories, limiting the use of each animal to once every 3 mo. Animals were examined by the veterinary staff after intubations to assess for trauma to upper airway tissue. Each examination was given a trauma grade of 0 for no visible signs of trauma, 1 if erythema of the larynx was present, 2 if visible excoriation of the mucus membranes was present, and 3 if bleeding (frank hemorrhage) was observed. The number of intubation attempts was restricted to 10 per animal per training session. A total of 170 intubations were completed on the ferrets during a 12-mo period. The average number of intubations per laboratory was 8.1 intubations per ferret. In addition, 1.8% of the intubations resulted in erythema (score, 1) after training, and 0.6% of the intubations resulted in excoriation (score, 2). Frank hemorrhage (score, 3) was not noted. The overall percentage of intubations resulting in any trauma during a training session was 0.02%. None of the animals have experienced any major complications to date. This ongoing training program has been used to teach neonatal intubation skills to emergency medicine residents for the past 12 mo. Ensuring the health and safety of the ferrets was paramount. Our results suggest that as many as 10 intubation attempts per session can be performed safely on each ferret without causing excessive trauma.