Conference Agenda

Session
Biomedical and Applications
Time:
Wednesday, 22/June/2022:
10:00am - 12:00pm

Session Chair: Ram Anand Vadlamani, Virginia Tech
Location: 301B

Oral Session

Presentations
10:00am - 10:20am
BioMed and Applications: 1

Pulsed Electric Field Activation of Platelet Rich Plasmas with Different Levels of Platelet Enrichment and Red Blood Cell Content

B. Neculaes1, A. Garner2, E. Longman2

1GE Research, Niskayuna, NY, United States of America; 2Purdue University, West Lafayette, IN, United States of America

Ex vivo platelet activation is being explored for a variety of regenerative medicine and wound healing applications. Activating the platelets releases multiple proteins and growth factors with key roles in the wound healing cascade. Platelet rich plasma (PRP) is typically obtained by centrifugation from the whole blood drawn from the patient. Bovine thrombin, a biochemical agent derived from animal sources, is then used in the clinic as the state of art platelet activator. The activated PRP, called platelet gel, is applied topically on the wound to be treated. Several studies have examined pulsed electric fields as a physically based method for platelet activation with several key advantages over bovine thrombin, including an easier workflow, the ability to standardize the method, reduced cost, enhanced tunability, and independence from animal sources that may cause immune response. Pulsed electric fields of various pulse durations [1] and delivery modality (capacitive or conductive coupling; various concentrations of extracellular calcium) [2] have been successfully used for ex vivo platelet activation in several biological matrices, including whole blood [3] and platelet rich plasma with red blood cell (RBC) content. This study presents the first results of pulsed electric field activation using three types of platelet rich plasmas from four human donors – one with RBC content and two others with minimal RBC content. Two types of microsecond pulsed electric fields have been used in this study, along with negative (no activation) and positive (activation using bovine thrombin) controls. Experimental results confirm growth factor release with pulsed electric fields for all three types of PRP – opening the door for wide clinical adoption of this novel pulsed power based biomedical approach.

[1] A. L. Garner, A. S. Torres, S. Klopman, and B. Neculaes, “Electrical stimulation of whole blood for growth factor release and potential clinical implications,” Med. Hypotheses, vol. 143, 2020, Art. no. 110105.

[2] A. L. Garner, A. L. Frelinger III, A. J. Gerrits, T. Gremmel, E. E. Forde, S. L. Carmichael, A. D. Michelson, and V. B. Neculaes, “Using extracellular calcium concentration and electric pulse conditions to tune platelet-rich plasma growth factor release and clotting,” Med. Hypotheses, vol. 125, pp. 100-105, 2019.

[3] B. Neculaes, A. L. Garner, S. Klopman, C. Morton, and A. S. Torres, “A multi-donor ex vivo platelet activation and growth factor release study using electric pulses with durations up to 100 microseconds,” IEEE Access, vol. 9, pp. 31340 – 31349, 2021.



10:20am - 10:40am
BioMed and Applications: 2

Analysis of the Role of Cellular Heating in Microsecond Irreversible Electroporation

W. J. Milestone1, Q. Hu2, A. M. Loveless3, A. L. Garner3, R. P. Joshi1

1Texas Tech University, Lubbock, TX 79409, USA; 2Eastern Michigan University, Ypsilanti, MI 48197, USA; 3Purdue University, West Lafayette, IN 47906, USA.

Irreversible electroporation (IRE) involving tumor ablation presents a minimally invasive treatment and has found a niche in oncological applications. It has proven to be a safe and effective procedure for treating many unresectable tumors. However, the use of a series of high frequency sinusoidal bipolar electric pulses, in the context of cellular drug delivery and/or irreversible electroporation, has not been studied to the best of our knowledge. This scheme, similar to the High Frequency Irreversible Electroporation (HFIRE) protocol, could prove to be of utility and synergistic effects of local membrane heating might well be beneficial in the context of this long wavetrain. In the present simulation study, two aspects of interest will be probed: (i) the role of cell heating in possibly promoting the successful uptake of drugs for treatment, and (ii) the possible synergistic interplay between the electric field and local membrane heating in reducing the required electroporation threshold.

In this work, membrane electroporation will be simulated based on a Smoluchowski continuum analysis discussed elsewhere by our group, together with spatio-temporal heating due to the power dissipation from the external bi-phasic source. We will consider two-dimensional transient heat flow with azimuthal symmetry in a single spherical cell. The following aspects will be analyzed and discussed in this presentation: (i) Changes induced by including heating, especially effects on pore formation dynamics. (ii) Quantitative assessment of the magnitude of heating caused by the applied electric fields and its dependence on wavetrain and field characteristics. This could define safe-operating limits and/or provide guidance towards optimum parameter space. Given that the vast parameter space depends on multiple factors, including cell size, pulse characteristics, electrical and thermal parameters of the biological system, applied waveforms, and number of pulses, only a few test cases will be probed. (iii) The present simulations will allow predictions and mechanistic insights into the level of electric field threshold reductions possible due to synergistic heating. (iv) And finally, the possibility of establishing large thermal gradients at the membrane for thermo-diffusive transport will also be quantitatively assessed.



10:40am - 11:20am
BioMed and Applications: 3

Enhanced inactivation of Gram-negative bacteria using Gram-positive antibiotics and nanosecond electric pulses

R. A. Vadlamani1, A. Dhanabal2, D. A Detwiler3, R. Pal1,2, J. McCarthy3, M. N Seleem1,2, A. L Garner2

1Virginia Tech; 2Purdue University; 3Nanovis

Physically disrupting microorganism membranes with electric pulses renders the resistance mechanisms that inhibit or excrete antibiotics inert, reducing the antibiotic dosages required and making ineffective antibiotics effective. The growing threat of antibiotic resistant infections combined with a lack of drugs in the discovery pipeline necessitates novel ways for enhancing existing antibiotic effectiveness [1]. Nanosecond electric pulses (NSEPs) can make Gram-positive antibiotics, which are abundant, effective against Gram-negative resistant strains of bacteria, for which new and effective medicines are sorely lacking, on a sufficiently short timeframe to prevent resistance mechanisms from developing. We demonstrate the synergistic inactivation of a Gram-positive (Staphylococcus aureus) and two Gram-negative (Escherichia coli, and Pseudomonas aeruginosa) bacteria by combining various antibiotics with different mechanisms of action with 222 30 kV/cm or 500 20 kV/cm, 300 ns duration electric pulses (EPs) [2], energy matched [3] but selected such that the lower electric fields had minimal impact on viability, but reacted synergistically in combination with antibiotics. Combining NSEPs with antibiotics induced several log-reduction of colony forming units for antibiotics that induced no inactivation following 10 minutes of exposure in solution without NSEPs. Staphylococcus aureus inactivation improved compared to EPs alone when we combined 2 mg/L or 20 mg/mL of rifampicin with the 30 kV/cm EPs; however, only a few of the other combinations enhanced inactivation. E. coli inactivation improved compared to EPs alone by combining either EP pulse train with 2 mg/L or 20 mg/mL of mupirocin or rifampicin or by combining the 30 kV/cm EPs with either 2 mg/L or 20 mg/mL of erythromycin or vancomycin. These results indicate that EPs can make Gram-positive antibiotics efficient for inactivating Gram-negative bacteria.

[1] D. S. Davies and E. S. Verde, “Antimicrobial resistance,” Search Collab. Solut. World Innov. Summit Health Doha, pp. 1–36, 2013.

[2] A. Vadlamani, D. A. Detwiler, A. Dhanabal, and A. L. Garner, “Synergistic bacterial inactivation by combining antibiotics with nanosecond electric pulses,” Appl. Microbiol. Biotechnol., vol. 102, no. 17, pp. 7589–7596, 2018.

[3] K. Schoenbach, R. Joshi, S. Beebe, and C. Baum, “A scaling law for membrane permeabilization with nanopulses,” IEEE Trans. Dielectr. Electr. Insul., vol. 16, no. 5, pp. 1224–1235, Oct. 2009.



11:20am - 11:40am
BioMed and Applications: 4

Using intense pulsed electric fields for the sterilization of solid pre-packed food – the design and preliminary testing of a practical MV-class system

M. Woodyard, B. Novac, P. Senior, J. Stobbs

Loughborough University, United Kingdom

A pulsed power MV-class system for generating intense pulsed electric fields in a very large volume of water was designed, manufactured and tested. This is a first and essential step towards the proof-of-principle demonstration of a novel technique for the non-invasive sterilisation of pre-packed food. The pulsed power system, based on MV Tesla transformer technology, is capable of producing pulsed electric fields in excess of 100 kV/cm in a massive volume (≈1 L) of water. The electric field distribution in the processing volume is obtained using a commercially available electrostatic solver, benchmarked using an electro-optic Kerr-effect assembly. The paper presents a detailed analysis of the unique processing unit and theoretically explores a possible scenario related to processing pre-packed food.



11:40am - 12:00pm
BioMed and Applications: 5

Resonant Charging Circuit for a Semiconductor-based Marx Generator for an Electropoation Device

M. Sack, D. Herzog, J. Ruf, G. Mueller

Karlsruhe Institute of Technology, Germany

Electroporation devices for the treatment of plant material in a continuous flow may employ a Marx circuit featuring parallel charging and a complete discharge of the capacitors during each pulse. Thereby, the use of semiconductor switches rather than spark gap switches allows for a modern design without the need of recurring maintenance of the spark gap switches. However, the use of semiconductor switches demands for an adaptation of the circuit to the properties of these switches being able to operate at lower voltage and current but higher pulse repetition rate compared to spark gap switches. For such a design, resonant charging turned out to be of advantage because it combines a fast energy transfer from a DC-link capacitor as part of the power supply to the stage capacitors of a Marx generator with a charging circuit having no active switches. Moreover, it enables an increase in the charging voltage by a factor of approximately two with respect to the DC-link voltage.
An 8-stage Marx circuit with a stage voltage of 1 kV has been set up to study resonant charging. Its charging path has been equipped with current-compensated chokes having a low inductance during charging and a high inductance for transient insulation of the stages during the pulse generation. The charging circuit has been designed for a peak current of 70 A and a charging time of 800 µs. A separate inductance between the DC-link capacitor of the power supply and the generator serves as the inductive component for the resonance circuit. As pulse switches IGBTs have been employed. In the course of the charging process, they serve as opening switches allowing for fine tuning of the charging voltage in repetitive operation. The generator has been operated either with a ground connection at its negative output terminal or grounded at its center to deliver a ground-symmetric output voltage. In both cases the magnetic energy stored inside the current-compensated chokes has been either degraded or recycled to prevent the cores from saturation. In the latter case, measurements revealed oscillations in a resonant circuit comprising the stray capacitance of the current-compensated chokes. A series diode avoids these oscillations. The paper describes selected details of the circuit design and presents measurements of the charging process.