In this paper, the features of the intensity-based Doppler variance (IBDV) method were analyzed systemically with a flow phantom. The effects of beam scanning density, flow rate and the time interval between neighboring A-lines on the performance of this method were investigated. The IBDV method can be used to quantify the flow rate and its sensitivity can be improved by increasing the time interval between the neighboring A-lines. A higher sensitivity IBDV method that applies the algorithm along the slower scan direction was proposed. In comparison to laser speckle imaging maps of blood flow, we demonstrated the ability of the method to identify vessels with altered blood flow. In clinical measurements, we demonstrated the ability of the method to image vascular networks with exquisite spatial resolution and at depths up to 1.2 mm in human skin. These results collectively demonstrated the potential of the method to monitor the microvasculature during disease progression and in response to therapeutic intervention.

Background/objectives

Current laser therapy of port wine stain (PWS) birthmarks with a single laser pulse (SLP) does not produce complete lesion removal in the majority of patients. To improve PWS therapeutic efficacy, we evaluated the performance of an approach based on multiple laser pulses (MLP) to enhance blood vessel photocoagulation.

Study design

The hamster dorsal window chamber model was used. Radiant exposure (RE), pulse repetition rate (f(r)), total number of pulses (n(p)), and length of vessel irradiated were varied. Blood vessels in the window were irradiated with either SLP with RE of 4-7 J/cm(2) or MLP with RE per pulse of 1.4-5.0 J/cm(2), f(r) of 0.5-26.0 Hz, and n(p) of 2-5. The laser wavelength was 532 nm and pulse duration was 1 ms. Either a 2 mm vessel segment or entire vessel branch was irradiated. Digital photographs and laser speckle images of the window were recorded before and at specific time points after laser irradiation to monitor laser-induced blood vessel structural and functional changes, respectively.

Results

We found that: (1) for a SLP approach, the RE required to induce blood vessel photocoagulation was 7 J/cm(2) as compared to only 2 J/cm(2) per pulse for the MLP approach; (2) for MLP, two pulses at a repetition rate of 5 Hz and a RE of 3 J/cm(2) can induce photocoagulation of more than 80% of irradiated blood vessel; and (3) irradiation of a longer segment of blood vessel resulted in lower reperfusion rate.

Conclusions

The MLP approach can induce blood vessel photocoagulation at much lower RE per pulse as compared to SLP. The 5 Hz f(r) and the need for two pulses are achievable with modern laser technology, which makes the MLP approach practical in the clinical management of PWS birthmarks.

BACKGROUND: Port-wine stain (PWS) is a congenital, progressive vascular malformation but the pathogenesis remains incompletely understood. OBJECTIVE: We sought to investigate the activation status of various kinases, including extracellular signal-regulated kinase, c-Jun N-terminal kinase, AKT, phosphatidylinositol 3-kinase, P70 ribosomal S6 kinase, and phosphoinositide phospholipase C γ subunit, in PWS biopsy tissues. METHODS: Immunohistochemistry was performed on 19 skin biopsy samples from 11 patients with PWS. RESULTS: c-Jun N-terminal kinase, extracellular signal-regulated kinase, and P70 ribosomal S6 kinase in pediatric and adult PWS blood vessels were consecutively activated. Activation of AKT and phosphatidylinositol 3-kinase was found in many adult hypertrophic PWS blood vessels but not in infants. Phosphoinositide phospholipase C γ subunit showed strong activation in nodular PWS blood vessels. LIMITATION: Infantile PWS sample size was small. CONCLUSION: Our data suggest a subsequent activation profile of various kinases during different stages of PWS: (1) c-Jun N-terminal and extracellular signal-regulated kinases are firstly and consecutively activated in all PWS tissues, which may contribute to both the pathogenesis and progressive development of PWS; (2) AKT and phosphatidylinositol 3-kinase are subsequently activated, and are involved in the hypertrophic development of PWS blood vessels; and (3) phosphoinositide phospholipase C γ subunit is activated in the most advanced stage of PWS and may participate in nodular formation.

Laser cartilage reshaping (LCR) with cryogen spray cooling is a promising modality for producing cartilage shape change while reducing cutaneous thermal injury. However, LCR in thicker tissues, such as auricular cartilage, requires higher laser power, thus increasing cooling requirements. To eliminate the risks of freeze injury characteristic of high cryogen spray pulse rates, a carbon dioxide (CO2) spray, which evaporates rapidly from the skin, has been proposed as the cooling medium. This study aims to identify parameter sets which produce clinically significant reshaping while producing minimal skin thermal injury in LCR with CO2 spray cooling in ex vivo rabbit auricular cartilage. Excised whole rabbit ears were mechanically deformed around a cylindrical jig and irradiated with a 1.45-μm wavelength diode laser (fluence 12-14 J/cm(2) per pulse, four to six pulse cycles per irradiation site, five to six irradiation sites per row for four rows on each sample) with concomitant application of CO2 spray (pulse duration 33-85 ms) to the skin surface. Bend angle measurements were performed before and after irradiation, and the change quantified. Surface temperature distributions were measured during irradiation/cooling. Maximum skin surface temperature ranged between 49.0 to 97.6 °C following four heating/cooling cycles. Significant reshaping was achieved with all laser dosimetry values with a 50-70 °C difference noted between controls (no cooling) and irradiated ears. Increasing cooling pulse duration yielded progressively improved gross skin protection during irradiation. CO2 spray cooling may potentially serve as an alternative to traditional cryogen spray cooling in LCR and may be the preferred cooling medium for thicker tissues. Future studies evaluating preclinical efficacy in an in vivo rabbit model are in progress.

Background and objectives

Complete blanching of port wine stain (PWS) birthmarks after laser therapy is rarely achieved for most patients. We postulate that the low therapeutic efficacy or treatment failure is caused by regeneration and revascularization of photocoagulated blood vessels due to angiogenesis associated with the skin's normal wound healing response. Rapamycin (RPM), an antiangiogenic agent, has been demonstrated to inhibit growth of pathological blood vessels. Our objectives were to (1) investigate whether topical RPM can inhibit reperfusion of photocoagulated blood vessels in an animal model and (2) determine the effective RPM concentration required to achieve this objective.

Study design/materials and methods

For both laser-only and combined laser and RPM treated animals, blood vessels in the dorsal window chambers implanted on golden Syrian hamsters were photocoagulated with laser pulses. Structural and flow dynamics of blood vessels were documented with color digital photography and laser speckle imaging to evaluate photocoagulation and reperfusion. For the combined treatment group, topical RPM was applied to the epidermal side of the window daily for 14 days after laser exposure.

Results

In the laser-only group, 23 out of 24 photocoagulated blood vessels reperfused within 5-14 days. In the combined treatment group with different RPM formulae and concentrations, the overall reperfusion rate of 36% was much lower as compared to the laser-only group. We also found that the reperfusion rate was not linearly proportional to the RPM concentration.

Conclusions

With topical RPM application, the frequency of vessel reperfusion was considerably reduced, which implies that combined light and topical antiangiogenic therapy might be a promising approach to improve the treatment efficacy of PWS birthmarks.

BACKGROUND: Administration of topical rapamycin (RPM) suppresses the regeneration and revascularization of photocoagulated blood vessels induced by pulsed dye laser (PDL). OBJECTIVE: To systematically elucidate the molecular pathophysiology of the inhibition of PDL-induced angiogenesis by topical RPM in a rodent model. METHODS: The mRNA expression profiles of 86 angiogenic genes and phosphorylation levels of ribosomal protein S6 kinase (P70S6K) in rodent skin were examined with or without topical RPM administration post-PDL exposure. RESULTS: The PDL-induced systematic increases in transcriptional levels of angiogenic genes showed a peak expression at days 3-7 post-PDL in rodent skin. Topical application of 1% RPM significantly and systematically suppressed the PDL-induced increase in mRNA levels of the examined angiogenic genes during the first five days post-PDL. The phosphorylation levels of P70S6K increased after PDL exposure but those increases were suppressed by the topical RPM. After topical application, RPM penetrated to an approximate depth of 768.4 μm into rodent skin. CONCLUSION: Topical application of 1% RPM can significantly and systematically suppress the PDL-induced early stage of angiogenesis via inhibition of the AKT/mTOR/P70S6K pathway in a rodent model.

Background/objectives

Similar to conventional cryogen spray cooling, carbon dioxide (CO2) spray may be used in combination with laser cartilage reshaping (LCR) to produce cartilage shape change while minimizing cutaneous thermal injury. Recent ex vivo evaluation of LCR with CO2 cooling in a rabbit model has identified a promising initial parameter space for in vivo safety and efficacy evaluation. This pilot study aimed to evaluate shape change and cutaneous injury following LCR with CO2 cooling in 5 live rabbits.

Study design/materials and methods

The midportion of live rabbit ears were irradiated with a 1.45 µm wavelength diode laser (12 J/cm(2)) with simultaneous CO2 spray cooling (85 millisecond duration, 4 alternating heating/cooling cycles per site, 5 to 6 irradiation sites per row for 3 rows per ear). Experimental and control ears (no LCR) were splinted in the flexed position for 30 days following exposure. A total of 5 ears each were allocated to the experimental and control groups.

Results

Shape change was observed in all irradiated ears (mean 70 ± 3°), which was statistically different from control (mean 37 ± 11°, P = 0.009). No significant thermal cutaneous injury was observed, with preservation of the full thickness of skin, microvasculature, and adnexal structures. Confocal microscopy and histology demonstrated an intact and viable chondrocyte population surrounding irradiated sites.

Conclusions

LCR with CO2 spray cooling can produce clinically significant shape change in the rabbit auricle while minimizing thermal cutaneous and cartilaginous injury and frostbite. This pilot study lends support for the potential use of CO2 spray as an adjunct to existing thermal-based cartilage reshaping modalities. An in vivo systematic evaluation of optimal laser dosimetry and cooling parameters is required.

Erythrocyte-based nanoparticle platforms can offer long circulation times not offered by traditional drug delivery methods. We have developed a novel erythrocyte-based nanoparticle doped with indocyanine green (ICG), the only FDA-approved near-infrared chromophore. Here, we report on the absorption and fluorescence emission characteristics of these nanoparticles fabricated using ICG concentrations in the range of 161-323 μM. These nanoparticles may serve as biocompatible optical materials for various clinical imaging and phototherapeutic applications.