We developed a novel localized antivascular method, namely photo-mediated ultrasound therapy (PUT), by applying synchronized laser and ultrasound pulses. PUT relies on high optical contrast among biological tissues. Taking advantage of the high optical absorption of hemoglobin, PUT can selectively target microvessels without causing unwanted damages to the surrounding tissue. Moreover, PUT working at different optical wavelengths can selectively treat veins or arteries by utilizing the contrast in the optical spectra between deoxy- and oxy-hemoglobin. Through our experiments, we demonstrated that cavitation might have played a key role in PUT. The addition of a laser pulse to an existing ultrasound field can significantly improve the likelihood of inertial cavitation, which can induce microvessel damage through its mechanical effect. In comparison with conventional laser therapies, such as photothermolysis and photocoagulation, the laser energy level needed in PUT is significantly lower. When a nanosecond laser was used, our in vivo experiments showed that the needed laser fluence was in the range of 4 to 40 mJ/cm2.
Optical coherence tomography (OCT) is a noninvasive interferometric imaging modality providing anatomical information at depths of millimeters and a resolution of micrometers. Conventional OCT images limit our knowledge to anatomical structures alone, without any contrast enhancement. Therefore, here we have, for the first time, optimized an OCT-based contrast-enhanced imaging system for imaging single cells and blood vessels in vivo inside the living mouse retina at subnanomolar sensitivity. We used bioconjugated gold nanorods (GNRs) as exogenous OCT contrast agents. Specifically, we used anti-mouse CD45 coated GNRs to label mouse leukocytes and mPEG-coated GNRs to determine sensitivity of GNR detection in vivo inside mice retinae. We corroborated OCT observations with hyperspectral dark-field microscopy of formalin-fixed histological sections. Our results show that mouse leukocytes that otherwise do not produce OCT contrast can be labeled with GNRs leading to significant OCT intensity equivalent to a 0.5 nM GNR solution. Furthermore, GNRs injected intravenously can be detected inside retinal blood vessels at a sensitivity of ∼0.5 nM, and GNR-labeled cells injected intravenously can be detected inside retinal capillaries by enhanced OCT contrast. We envision the unprecedented resolution and sensitivity of functionalized GNRs coupled with OCT to be adopted for longitudinal studies of retinal disorders.
Application of femtosecond lasers to cataract surgery has added unprecedented precision and reproducibility but ocular safety limits for the procedure are not well-quantified. We present an analysis of safety during laser cataract surgery considering scanned patterns, reduced blood perfusion, and light scattering on residual bubbles formed during laser cutting. Experimental results for continuous-wave 1030 nm irradiation of the retina in rabbits are used to calibrate damage threshold temperatures and perfusion rate for our computational model of ocular heating. Using conservative estimates for each safety factor, we compute the limits of the laser settings for cataract surgery that optimize procedure speed within the limits of retinal safety.
Femtosecond lasers have added unprecedented precision and reproducibility to cataract surgery. However, retinal safety limits for the near-infrared lasers employed in surgery are not well quantified. We determined retinal injury thresholds for scanning patterns while considering the effects of reduced blood perfusion from rising intraocular pressure and retinal protection from light scattering on bubbles and tissue fragments produced by laser cutting. We measured retinal damage thresholds of a stationary, 1030-nm, continuous-wave laser with 2.6-mm retinal spot size for 10- and 100-s exposures in rabbits to be 1.35 W (1.26 to 1.42) and 0.78 W (0.73 to 0.83), respectively, and 1.08 W (0.96 to 1.11) and 0.36 W (0.33 to 0.41) when retinal perfusion is blocked. These thresholds were input into a computational model of ocular heating to calculate damage threshold temperatures. By requiring the tissue temperature to remain below the damage threshold temperatures determined in stationary beam experiments, one can calculate conservative damage thresholds for cataract surgery patterns. Light scattering on microbubbles and tissue fragments decreased the transmitted power by 88% within a 12 deg angle, adding a significant margin for retinal safety. These results can be used for assessment of the maximum permissible exposure during laser cataract surgery under various assumptions of blood perfusion, treatment duration, and scanning patterns.
Decreasing the pulse duration helps confine damage, shorten treatment time, and minimize pain during retinal photocoagulation. However, the safe therapeutic window (TW), the ratio of threshold powers for thermomechanical rupture of Bruch's membrane and mild coagulation, also decreases with shorter exposures. Two potential approaches toward increasing TW are investigated: (a) decreasing the central irradiance of the laser beam and (b) temporally modulating the pulse. An annular beam with adjustable central irradiance was created by coupling a 532-nm laser into a 200-μm core multimode optical fiber at a 4-7 deg angle to normal incidence. Pulse shapes were optimized using a computational model, and a waveform generator was used to drive a PASCAL photocoagulator (532 nm), producing modulated laser pulses. Acute thresholds for mild coagulation and rupture were measured in Dutch-Belted rabbit in vivo with an annular beam (154-163 μm retinal diameter) and modulated pulse (132 μm, uniform irradiance "flat-top" beam) with 2-50 ms pulse durations. Thresholds with conventional constant-power pulse and a flat-top beam were also determined. Both annular beam and modulated pulse provided a 28% increase in TW at 10-ms duration, affording the same TW as 20-ms pulses with conventional parameters.
This study evaluates the effects of exposure duration, beam diameter, and power on the safety, selectivity, and healing of
retinal lesions created using a continuous line scanning laser. A 532 nm laser (PASCALTM) with retinal beam diameters
of 40 and 66 μm was applied to 60 eyes of 30 Dutch-Belted rabbits. Retinal exposure duration varied from 15 to 60 μs.
Lesions were acutely assessed by ophthalmoscopy and fluorescein angiography (FA). RPE flatmounts were evaluated
with live-dead fluorescent assay (LD). Histological analysis was performed at 1 hour, 1 and 3 days, 1 and 2 weeks, and 1
and 2 months following laser treatment. Ophthalmoscopic visibility (OV) of the lesions corresponded to photoreceptor
damage on histological analysis at 1 hour. In subvisible lesions, FA and LD yielded similar thresholds of RPE damage.
The ratios of the threshold of rupture and of OV to FA visibility (measures of safety and selectivity) increased with
decreasing duration and beam diameter. Above the threshold of OV, histology showed focal RPE damage and
photoreceptor loss at one day without inner retinal effects. By one week, continuity of photoreceptor and RPE layers was
restored. By 1 month, photoreceptors appeared normal while hypertrophy and hyperpigmentation of the RPE persisted.
Retinal therapy with a fast scanning continuous laser achieves selective targeting of the RPE and, at higher power, of the
photoreceptors. The damage zone in the photoreceptor layer is quickly filled-in, likely due to photoreceptor migration
from adjacent zones. Continuous scanning laser can treat large retinal areas within standard eye fixation time.
Shorter pulse durations help confine thermal damage during retinal photocoagulation, decrease treatment time and minimize pain. However, safe therapeutic window (the ratio of threshold powers for rupture and mild coagulation) decreases with shorter exposures. A ring-shaped beam enables safer photocoagulation than conventional beams by reducing the maximum temperature in the center of the spot. Similarly, a temporal pulse modulation decreasing its power over time improves safety by maintaining constant temperature for a significant portion of the pulse. Optimization of the beam and pulse shapes was performed using a computational model. In vivo experiments were performed to verify the predicted improvement. With each of these approaches, the pulse duration can be decreased by a factor of two, from 20 ms down to 10 ms while maintaining the same therapeutic window.
In laser retinal photocoagulation, short (<20 ms) pulses have been found to reduce thermal damage to the inner retina, decrease treatment time, and minimize pain. However, the safe therapeutic window (defined as the ratio of power for producing a rupture to that of mild coagulation) decreases with shorter exposures. To quantify the extent of retinal heating and maximize the therapeutic window, a computational model of millisecond retinal photocoagulation and rupture was developed. Optical attenuation of 532-nm laser light in ocular tissues was measured, including retinal pigment epithelial (RPE) pigmentation and cell-size variability. Threshold powers for vaporization and RPE damage were measured with pulse durations ranging from 1 to 200 ms. A finite element model of retinal heating inferred that vaporization (rupture) takes place at 180-190°C. RPE damage was accurately described by the Arrhenius model with activation energy of 340 kJ/mol. Computed photocoagulation lesion width increased logarithmically with pulse duration, in agreement with histological findings. The model will allow for the optimization of beam parameters to increase the width of the therapeutic window for short exposures.
Short duration (< 20 ms) pulses are desirable in patterned scanning laser photocoagulation to confine thermal damage to
the photoreceptor layer, decrease overall treatment time and reduce pain. However, short exposures have a smaller
therapeutic window (defined as the ratio of rupture threshold power to that of light coagulation). We have constructed a
finite-element computational model of retinal photocoagulation to predict spatial damage and improve the therapeutic
window. Model parameters were inferred from experimentally measured absorption characteristics of ocular tissues, as
well as the thresholds of vaporization, coagulation, and retinal pigment epithelial (RPE) damage. Calculated lesion
diameters showed good agreement with histological measurements over a wide range of pulse durations and powers.
In patterned scanning laser photocoagulation, shorter duration (< 20 ms) pulses help reduce thermal
damage beyond the photoreceptor layer, decrease treatment time and minimize pain. However, safe
therapeutic window (defined as the ratio of rupture threshold power to that of light coagulation) decreases
for shorter exposures. To quantify the extent of thermal damage in the retina, and maximize the therapeutic
window, we developed a computational model of retinal photocoagulation and rupture. Model parameters
were adjusted to match measured thresholds of vaporization, coagulation, and retinal pigment epithelial
(RPE) damage. Computed lesion width agreed with histological measurements in a wide range of pulse
durations and power. Application of ring-shaped beam profile was predicted to double the therapeutic
window width for exposures in the range of 1 - 10 ms.
Pan-retinal photocoagulation in patients with diabetic retinopathy typically involves application of more than
1000 laser spots; often resulting in physician fatigue and patient discomfort. We present a semi-automated patterned
scanning laser photocoagulator that rapidly applies predetermined patterns of lesions; thus, greatly improving the
comfort, efficiency and precision of the treatment.
Patterns selected from a graphical user interface are displayed on the retina with an aiming beam, and
treatment can be initiated and interrupted by depressing a foot pedal. To deliver a significant number of burns during
the eye's fixation time, each pulse should be considerably shorter than conventional 100ms pulse duration. We
measured coagulation thresholds and studied clinical and histological outcomes of the application of laser pulses in the
range of 1-200ms in pigmented rabbits.
Laser power required for producing ophthalmoscopically visible lesions with a laser spot of 132&mgr;m decreased
from 360 to 37mW with pulse durations increasing from 1 to 100ms. In the range of 10-100ms clinically and
histologically equivalent light burns could be produced. The safe therapeutic range of coagulation (ratio of the laser
power required to produce a rupture to that for a light burn) decreased with decreasing pulse duration: from 3.8 at 100ms, to 3.0 at 20ms, to 2.5 at 10ms, and to 1.1 at 1ms. Histology demonstrated increased confinement of the thermal damage with shorter pulses, with coagulation zone limited to the photoreceptor layer at pulses shorter than 10ms. Durations of 10-20ms appear to be a good compromise between the speed and safety of retinal coagulation. Rapid application of multiple lesions greatly improves the speed, precision, and reduces pain in retinal photocoagulation.