In-depth analysis of antibacterial mechanisms of laser generated shockwave treatment.

Background and Objectives Laser generated shockwave (LGS) is a novel modality for minimally invasive disruption of bacterial biofilms. The objectives of this study are to determine the mechanisms behind LGS treatment and non-biofilm effects on bacterial disruption, including (1) comparing bacterial load with and without LGS in its planktonic form and (2) estimating bacterial cell permeability following LGS. Study Design/Materials and Methods For the first study, planktonic S. epidermidis were treated with gentamicin (0, 8, 16, 32, 64 µg/ml) with and without LGS (1064 nm Nd:YAG laser, 110.14 mJ/mm2 , pulse duration 9 ns, spot size 3 mm, n = 8/group), and absorbances at 600 nm compared. For the second study, four samples of planktonic S. epidermidis were treated with LGS (same settings). Propidium iodide (PI) uptake via flow cytometry as a measure of cell permeability was measured at 0, 10, and 20 minutes following LGS. RESULTS: In comparing corresponding gentamicin concentrations within both LGS-treated samples and controls at 0 hours, there were no differences in absorbance (P = 0.923 and P = 0.814, respectively). Flow cytometry found modest PI uptake (10.4 ± 2.5%) immediately following LGS treatment, with time-dependent increase and persistence of the signal at 20 minutes (R2 = 0.449, P = 0.048). CONCLUSION: Taken together, LGS does not appear to have direct bacteriocidal properties, but rather by allowing for biofilm disruption and bacterial cell membrane permeabilization, both of which likely increase topical antibiotic delivery to pathogenic organisms. Insight into the mechanisms of LGS will allow for improved clinical applications and facilitate safe and effective translation of this technology. Lasers Surg. Med. © 2018 Wiley Periodicals, Inc.

Vibroacoustographic System for Tumor Identification.

Oral and head and neck squamous cell carcinoma (OSCC) is the sixth most common cancer worldwide. The primary management of OSCC relies on complete surgical resection of the tumor. Margin-free resection, however, is difficult given the devastating effects of aggressive surgery. Currently, surgeons determine where cuts are made by palpating edges of the tumor. Accuracy varies based on the surgeon's experience, the location and type of tumor, and the risk of damage to adjacent structures limiting resection margins. To fulfill this surgical need, we contrast tissue regions by identifying disparities in viscoelasticity by mixing two ultrasonic beams to produce a beat frequency, a technique termed vibroacoustography (VA). In our system, an extended focal length of the acoustic stress field yields surgeons' high resolution to detect focal lesions in deep tissue. VA offers 3D imaging by focusing its imaging plane at multiple axial cross-sections within tissue. Our efforts culminate in production of a mobile VA system generating image contrast between normal and abnormal tissue in minutes. We model the spatial direction of the generated acoustic field and generate images from tissue-mimicking phantoms and ex vivo specimens with squamous cell carcinoma of the tongue to qualitatively demonstrate the functionality of our system. These preliminary results warrant additional validation as we continue clinical trials of ex vivo tissue. This tool may prove especially useful for finding tumors that are deep within tissue and often missed by surgeons. The complete primary resection of tumors may reduce recurrence and ultimately improve patient outcomes.

Laser-generated shockwaves enhance antibacterial activity against biofilms in vitro.

BACKGROUND AND OBJECTIVES: Bacterial biofilm formation within chronic wound beds, which provides an effective barrier against antibiotics, is a known cause of recalcitrant infections and a significant healthcare burden, often requiring repeated surgical debridements. Laser-generated shockwaves (LGS) is a novel, minimally invasive, and nonthermal modality for biofilm mechanical debridement which utilizes compressive stress waves, generated by photonic absorption in thin titanium films to mechanically disrupt the biofilm. Prior studies have demonstrated LGS monotherapy to be selectively efficacious for biofilm disruption and safe for host tissues. In this study, we sought to determine if LGS can enhance the antimicrobial activity and biofilm disruption capability of topical antibiotic therapy. STUDY DESIGN/MATERIALS AND METHODS: Staphylococcus epidermidis biofilms grown in vitro on glass were treated with topical gentamicin (31, 62, and 124 µg/ml) with and without LGS (n = 3-11/treatment group). Mechanical shockwaves were generated with a 1,064 nm Nd:YAG laser (laser fluence 110.14 mJ/mm2 , pulse duration 5 ns, spot size 3 mm). Following a 24-hour incubation period, bacterial viability was assessed by determining the number of colony-forming units (CFU) via the Miles and Misra method. Residual biofilm bioburden was analyzed using the crystal violet biofilm assay. RESULTS: With gentamicin monotherapy, CFU density (CFU/mm2 ) at 31, 62, and 124 µg/ml were (282 ± 84) × 104 , (185 ± 34) × 104 , and (113 ± 9) × 104 , respectively. With LGS and gentamicin therapy, CFU density decreased to (170 ± 44) × 104 , (89 ± 24) × 104 , and (43 ± 3) × 104 , respectively (P = 0.1704, 0.0302, and 0.0004 when compared with gentamicin alone). Biofilm burden as measured by the assay in the gentamicin 31, 62, and 124 µg/ml groups was reduced by 80%, 95%, and 98% when LGS was added (P = 0.0102, >0.0001, and 0.0001 for all groups when compared with gentamicin alone). Furthermore, samples treated with LGS saw an increase in susceptibility to gentamicin, in terms of reduced biofilm bioburden and CFU densities. CONCLUSION: LGS enhances the efficacy of topical antibiotics in an in vitro model. This has significant implications for clinical applications in the management of chronic soft tissue infections and recalcitrant chronic rhinosinusitis. Lasers Surg. Med. 49:539-547, 2017. © 2017 Wiley Periodicals, Inc.

Analysis of flexible substrates for clinical translation of laser-generated shockwave therapy.

Bacteria biofilms in chronically infected wounds significantly increase the burden of healthcare costs and resources for patients and clinics. Because biofilms are such an effective barrier to standard antibiotic treatment, new methods of therapy need to be developed to combat these infections. Our group has demonstrated the potential of using Laser Generated Shockwaves as a potential therapy to mechanically disrupt the bacterial biofilms covering the wound. Previous studies have used rigid silica glass as the shockwave propagation medium, which is not compatible with the intended clinical application. This paper describes the exploration of five candidate flexible plastic films to replace the glass substrate. Each material measured 0.254 mm thick and was used to generate shockwaves of varying intensities. Shockwave characterization was performed using a high-speed Michelson displacement interferometer and peak stress values obtained in the flexible substrates were compared to glass using one-way nested Analysis of Variance and Tukey HSD post-hoc analysis. Results demonstrate statistically significant differences between substrate material and indicate that polycarbonate achieves the highest peak stress for a given laser fluence suggesting that it is optimal for clinical applications.