Misty L. Noble

Polyacrylamide gel as an acoustic coupling medium for focused ultrasound therapy

A hydrogel acoustic coupling medium was investigated as a practical alternative to water for clinical applications of focused ultrasound (US) therapy. Material characterization and functional testing of polyacrylamide gel couplers were performed. Acoustic, bulk and thermal properties were measured. Conical couplers were designed and fabricated to fit a 3.5-MHz, spherically concave transducer for functional tests, including Schlieren imaging, power efficiency measurements and in vivo hemostasis experiments. Polyacrylamide was shown to have favorable acoustic properties that varied linearly with acrylamide concentration from 10% to 20% weight in volume. Attenuation coefficient, sound speed and impedance ranged from 0.08 to 0.14 dB/cm at 1 MHz, 1546 to 1595 m/s and 1.58 to 1.68 Mrayl, respectively. An intraoperative in vivo hemostasis experiment in a sheep model demonstrated that the gel-coupled transducer was capable of inducing hemostasis in actively bleeding splenic and hepatic incisions. The results of this study show that polyacrylamide may be a promising coupling material for focused US therapy.

Sustained release of antibiotic from poly(2-hydroxyethyl methacrylate) to prevent blinding infections after cataract surgery.

Intraocular lens implantation after opacified natural lens removal is the primary treatment for cataracts in developed countries. Cataract surgery is generally considered safe, but entails significant risks in countries where sophisticated sterile operating theaters are not widely available. Post-operative infection (endophthalmitis) is a potential blinding complication. Infection often results from bacterial colonization of the new lens implant and subsequent antibiotic-tolerant biofilm formation. To combat this risk, we developed a polymeric hydrogel system that can deliver effective levels of antibiotic over an extended period of time within the globe of the eye. Norfloxacin antibiotic was loaded into cross-linked poly(2-hydroxyethyl methacrylate) (pHEMA) gels, which were subsequently surface-modified with octadecyl isocyanate to produce a hydrophobic rate-limiting barrier controlling norfloxacin release. Octadecyl surface modification was characterized using scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS). A 15-min modification leads to a uniform surface coating and near zero order release of norfloxacin from the matrix. Norfloxacin released from coated pHEMA kills Staphylococcus epidermidis in suspension and on a simulated medical implant surface. With these data, we demonstrate a new and effective system for sustained drug release from a hydrogel matrix with specific application for intraocular lens surgery.

Liver Hemostasis With High-Intensity Ultrasound Repair and Healing

Objective. Previous studies have shown that high‐intensity focused ultrasound can effectively control bleeding from injuries of liver, spleen, and blood vessels. This study investigated long‐term hemostasis and tissue repair after high‐intensity focused ultrasound treatment in liver. Methods. A total of 21 rabbits were randomly assigned to 2 groups: high‐intensity focused ultrasound treatment (n = 14) and sham treatment (n = 7). All animals had sterile laparotomy and liver exposure. The high‐intensity focused ultrasound–treated animals received liver incisions, 20 to 25 mm long and 4 to 6 mm deep, followed immediately by high‐intensity focused ultrasound application until complete hemostasis was achieved. After recovery, sonographic images, blood samples, and histologic samples were collected immediately and on days 1, 3, 7, 14, 28, and 60 after treatment. Results. All 14 liver injuries were hemostatic after an average ± SD of 78 ± 44 seconds of high‐intensity focused ultrasound application, with no rebleeding at any time point after the treatment. Subsequent blood analysis showed no significant difference in serial hematologic or coagulation measures between the high‐intensity focused ultrasound and sham groups. Alanine aminotransferase and aspartate aminotransferase levels increased immediately after surgery by as much as 285% up to day 3 and returned to normal values by day 7. Hematocrit and white blood cell counts showed no statistically significant difference from normal values at all time points. Histologic examination up to 60 days after treatment revealed scarring and liver tissue regeneration at the treatment site. Conclusions. High‐intensity focused ultrasound appears to provide long‐lasting hemostasis of acute liver injury. Healing and repair mechanisms after high‐intensity focused ultrasound application appear to be intact.

Development and characterization of an innovative synthetic tissue-mimicking material for high intensity focused ultrasound (HIFU) exposures

While many tissue-mimicking phantoms have been developed for ultrasound imaging applications, none is suitable for exploration of the high temperature and pressure regimes involved in High Intensity Focused Ultrasound (HIFU). HIFU dosimetry studies are usually performed on biological tissues, but this approach has two drawbacks: 1) tissues are opaque and development of coagulative lesions cannot be visually observed in real-time, and 2) the natural heterogeneous structure of tissue may complicate direct comparison with numerical models. To address these issues, a new optically transparent tissue phantom was developed. It is a polyacrylamide hydrogel with a thermally sensitive indicator protein (Bovine Serum Albumin, 3 - 9%) that becomes optically diffusive when denatured. We describe various measurements undertaken to characterize this material and to demonstrate how well it matches tissue in terms of bulk acoustic and thermal properties. In summary, this new phantom material simulates many of the lesion-forming characteristics of soft tissue under HIFU exposures.

Sustained antibiotic release from an intraocular lens-hydrogel assembly for cataract surgery.

PURPOSE To develop a simple, novel polymeric drug-delivery device for prevention of postoperative bacterial infection after cataract surgery in the developing world. METHODS A poly(2-hydroxyethyl-methacrylate) (pHEMA) hydrogel was developed to achieve sustained release characteristics of antibiotics. The in vitro antibiotic release kinetics and efficacy of antibiotic function were tested using a silicone biofilm model. In vivo feasibility was investigated using a rabbit model. The control group of rabbits underwent standard cataract surgery with intraocular lens (IOL) implant and postoperative topical antibiotic and steroid. The experimental group received the polymeric device inserted with standard three-piece IOL at the time of surgery and received only topical steroids postoperatively. In vivo intraocular antibiotic levels and outcomes after cataract surgery were evaluated. RESULTS The in vitro studies demonstrate the antibiotic release kinetics can be controlled by optimization of the surface coating. The in vivo results showed sustained sufficient antibiotic concentration (above minimum inhibitory concentration for most common bacteria related to endophthalmitis) for >4 weeks. There was minimum toxicity observed in vivo. The device was effective in treating induced intraocular infection after cataract surgery. CONCLUSIONS The initial findings of the polymeric drug-delivery device demonstrate the feasibility delivering sufficient antibiotic in the anterior chamber for the immediate postoperative period in a rabbit model. The device is simple to produce and may help alleviate the potential postsurgical infections in the developing nations.

Treatment of uterine fibroid tumors in an in situ rat model using high-intensity focused ultrasound.

OBJECTIVE To determine the efficacy and safety of high-intensity focused ultrasound (HIFU) for the treatment of uterine fibroid tumors in an in situ animal model. DESIGN High-intensity focused ultrasound was applied intraoperatively to uterine fibroid tumors in rats. SETTING Department of Bioengineering, and Applied Physics Laboratory, University of Washington, Seattle, Washington. ANIMAL(S) Thirty-five tumors in 27 Eker rats that had spontaneous in situ uterine fibroids were randomly assigned into two groups receiving HIFU (n = 29) or sham (n = 6) treatments. INTERVENTION(S) Animals were anesthetized, and tumors were exposed surgically. The HIFU was applied at 3.5 MHz in 10-second bursts to produce coagulative necrosis lesions (3 mm by 10 mm), spaced 5 mm apart. Sham treatments consisted of exposing the tumors, and handling them similarly to those in the HIFU treatment group, but HIFU was not applied. MAIN OUTCOME MEASURE(S) Tumor volume was measured every week transabdominally using B-mode ultrasound imaging. Gross examination and histological analysis were performed after euthanasia. RESULT(S) More than half of the tumors in the HIFU treatment group showed significant tumor volume reduction. The average tumor volume in the sham treatment group increased 40-fold. Gross and histological analysis showed coagulative necrosis of tumor cells in the HIFU treatment group. CONCLUSION(S) The HIFU may provide an effective and safe method of treating uterine fibroid tumors.

Digital drug delivery: on–off ultrasound controlled antibiotic release from coated matrices with negligible background leaching

Hydrogels, such as crosslinked poly(2-hydroxyethyl methacrylate) (pHEMA) have been used extensively in controlled release drug delivery systems. Our previous work demonstrated an ultrasound (US)-responsive system based on pHEMA coated with a self-assembled multilayer of C12-C18 methylene chains. The resulting coating was predominantly crystalline and relatively impermeable, forming an US-activated switch that controlled drug release on-demand, and kept the drug within the matrix in the absence of US. The device, as developed did, however, show a low background drug-leaching rate independent of US irradiation. For some applications, it is desirable to have very low or zero background release rates. This was achieved here by a combination of new processing steps, and by copolymerizing HEMA with a relatively hydrophobic monomer, hydroxypropyl methacrylate (HPMA). These advances produced systems with undetectable ciprofloxacin background release rates that are capable of US-facilitated drug release - up to 14-fold increases relative to controls both before and after US exposure. In addition, these observations are consistent with the hypothesis that US-mediated disorganization of the coating allows a transient flux of water into the matrix where its interaction with bound and dissolved drug facilitates its movement both within and out of the matrix.

Intra‐operative Hemostasis of Punctured Femoral Artery Using HIFU: A Survival Study

The objective was to investigate the long‐term efficacy of hemostasis and healing of arteries after HIFU application. The femoral arteries of 22 adult rabbits were surgically exposed. Fifteen arteries were punctured with a needle and treated with HIFU, and 7 arteries were sham‐treated (no puncture or HIFU was applied). The tip of the HIFU applicator was positioned on the bleeding site, and HIFU energy was applied until hemostasis was achieved. The focal intensity was approximately 3,000 W/cm2, at the resonant frequency of 9.6 MHz. Serial ultrasound images, blood and tissue samples were collected immediately and on days 1, 3, 7, 14, 28, and 60 after the treatment. Eleven of the arteries were patent after the treatment, and four arteries were occluded, as confirmed using Doppler imaging. One of the occluded arteries reopened at day 14. HIFU exposure time to achieve hemostasis was 27 ±17 seconds for patent arteries and 101±38 seconds for the occluded arteries. The blood flow velocities were not statistically...

Effective ultrasound-targeted microbubble destruction (UTMD)-mediated gene transfer into the livers of small and large animals

Ultrasound (US)-targeted microbubble (MB) destruction (UTMD) can significantly enhance gene delivery in mouse livers when pDNA/MBs were injected into the portal vein (PV) with simultaneous US exposure using a focused transducer. However, this transducer was ineffective in enhancing gene transfer into rats. A 13-mm diameter unfocused transducer was designed and the delivery route of pDNA/MBs was modified into a specific liver lobe, resulting in >100-fold increase in luciferase expression in rats. To facilitate the translation into human applications, many technical issues were explored in large animal models. We applied 1.1 MHz US to targeted canine and swine liver lobes with simultaneous injection of pDNA/MBs into a PV segmental branch and occlusion of the inferior vena cava. For more effective treatment of large tissue volumes, a 52-mm apodized, dual element unfocused transducer was specifically constructed to reduce the near field transaxial pressure variations, producing a uniform field of US exposure. Together with a 15 kW-capacity US amplifier, a 692-fold and 1800-fold increases of gene expression in canines and swines were achieved at 2.7-MPa, respectively. Transaminase levels and histology analysis indicated minimal tissue damage. These results demonstrated that UTMD is highly promising for safe and efficient gene delivery into the liver.

268. Enhanced Gene Expression of Factor VIII by Ultrasound-Mediated Gene Delivery in Dogs

Previously we demonstrated that ultrasound (US)-mediated gene delivery (UMGD) can significantly enhance reporter gene transfer into the mouse and rat livers. This nonviral gene transfer strategy can bypass many obstacles encountered by viral gene therapy. Most significantly, we have achieved therapeutic levels of FVIII following UMGD into hemophilia A mice. Recently we have successfully developed prototype US systems including several unfocused and semi-focused transducers to treat large tissue volumes in canine and swine. In order to facilitate the translation of this technology to treat hemophilia, we have recently treated 2 normal dogs with UMGD of FVIII plasmids using an open surgery procedure. Four mg of a high-expressing, liver-specific pBS-HCRHPI-FVIIIA plasmid and 3 ml of Definity® MBs in 8 ml total PBS solution were injected via the segmental portal vein branch with simultaneous exposure of the target liver lobe to therapeutic US (1.1 MHz frequency, 20 cycle pulses, 50 Hz pulse repetition frequency) for 4 minutes using the large diameter transducers. A sham-treated dog received an equivalent pGL4/MB dose, but was not exposed to tUS. We used an apodized dual element unfocused transducer H105 at 2.5Mpa peak negative pressure (PNP) in the first dog experiment and observed low levels of hFVIII gene expression in the treated liver lobes. The second dog experiment was performed using a cylindrically 6.2 MPa at the focal area. One day following treatment, the treated liver lobes were sectioned, and representative sections were fixed and stained for FVIII expression by histochemical staining using a polyclonal anti-FVIII antibody. Untreated normal dog liver and human liver were used as negative and positive controls. Significant hFVIII gene expression was obtained in both treated liver lobes with the expression levels slightly lower than control human liver lobe. Furthermore, fairly homogeneous distribution of FVIII gene expression was observed in hepatocytes. Next, we performed Western blot analysis to confirm if the staining is specific to human FVIII protein. A heavy chain band specific to hFVIII was observed in treated dog plasma and positive hFVIII control, but not in control normal dog plasma and pre-bleed plasma sample from the dog before treatment. Significant enhancement of FVIII-specific clotting activity was also obtained in the second treated dog compared to pre-treatment and the first treated dog. In addition, transaminase levels and histology analysis indicated minimal tissue damage in treated dog livers. Enhancement of FVIII gene expression by UMGD is currently evaluated in hemophilia dogs. Our results sugges that UMGD has great potential for therapeutic treatment of hemophilia A.