The importance of long-term monitoring to evaluate the microvascular response to light-based therapies.

TO THE EDITOR Optimization of laser therapy for disfiguring vascular birthmarks is one specific clinical application (Kelly et al., 2005). Current treatment protocols involve the use of high-power pulsed laser irradiation with parameters chosen to induce selective photocoagulation of the targeted blood vessels, a method known as selective photothermolysis (Anderson and Parrish, 1983). Protocol design is based largely on results from numerical modeling studies (van Gemert et al., 1997), which are designed to predict the laser light distribution within the skin and subsequent photothermal response leading toward selective photocoagulation. However, current modeling methods do not incorporate adequately the complex dynamics associated with changes in light absorption due to conversion of hemoglobin to methemoglobin (Barton et al., 2001; Kimel et al., 2005) and convective mixing of blood during pulsed laser irradiation (Kimel et al., 2003), limiting their overall predictive capability. Furthermore, these models do not consider the chronic, biological response of the microvasculature to therapeutic laser intervention, which remains a poorly researched field. Knowledge of the biological response is critical to understand the repair processes initiated with photothermal injury and to assess the ultimate efficacy of the treatment. Animal models used as a platform to study light-based, microvascular-targeted therapies include the chick chorioallantoic membrane (Kimel et al., 1994, 2003), hamster cheek pouch (Suthamjariya et al., 2004), and rodent dorsal window chamber (Barton et al., 1998, 1999, 2001; Choi et al., 2004; Babilas et al., 2005; Smith et al., 2006). Optical imaging modalities used to evaluate noninvasively therapeutic outcome include video imaging, fluorescence microscopy, Doppler optical coherence tomography, and laser speckle imaging. Typically, short-term (o24 hours after intervention) evaluation of the microvasculature is performed. Babilas et al. (2005) proposed that a (1) 24-hour monitoring period allows for evaluation of ‘‘delayed biological effects’’ in the microvascular response to pulsed laser irradiation, and (2) the short-term response correlates well with numerical modeling predictions of photocoagulation. Longer (424 hour after intervention) monitoring periods usually are not performed with nontumor-bearing window chambers, presumably due to the reduced clarity of the chamber imposed by poor maintenance of window integrity secondary to infection. However, we hypothesized that the short-term response of the microvasculature is a poor predictor of the long-term response. With emphasis on aseptic methods, we have been able to maintain clear window chamber preparations for as long as 45 days after intervention. With this model, we have studied the long-term microvascular response to light-based, microvasculartargeted therapies. The data presented herein were acquired from adult male Golden Syrian hamsters. The surgery was performed as defined in a protocol approved by the University of California, Irvine, Animal Use Committee. The surgical protocol was a modified version of one described previously (Papenfuss et al., 1979). For all steps, aseptic conditions were maintained. We used wide-field color reflectance imaging and laser speckle imaging (Choi et al., 2004, 2006; Smith et al., 2006) to document and evaluate quantitatively and chronically ensuing blood flow dynamics. In one set of experiments, we irradiated select arteriolevenule pairs with laser pulse sequences to evaluate the efficacy of various therapeutic protocols. In the presented example (Figure 1a), we irradiated an arteriole-venule pair (upper circle in ‘‘Before’’ image) with five laser pulses containing both 532 and 1064 nm laser wavelengths and a second pair (lower circle) with a single 532/1064 nm laser pulse. Numerical modeling data suggested that both sets of laser parameters should induce photocoagulation in the B Choi et al. Long-Term Evaluation of Microvascular Response