Membrane attack complex (MAC) deposits in skin are not always accompanied by S-protein and clusterin.

Clusterin and S-protein bind to the membrane attack complex of complement (MAC) rendering it cytolytically inactive. Tissue necrosis as produced by pulsed tunable dye laser therapy (PTDL), and immune complex-related diseases such as lupus erythematosus, are accompanied by local accumulation of MAC. However, the mechanisms responsible for this accumulation might differ, and lead to deposition of MAC in different forms (cytolytically active or inactive). Biopsy specimens of lesional (22) and non-lesional (10) skin from 27 patients with a positive lupus band test (LBT) were studied using monoclonal antibodies against clusterin, S-protein, and MAC by immunofluorescence and immunoperoxidase. Identical studies were performed in normal and angiomatous skin specimens from three normal individuals before and after laser irradiation. MAC was present in 30 of 32 positive LBT skin biopsies. MAC was not only present in lesional (21 of 22) but also in non-lesional skin (nine of 10), although the intensity of staining appeared to be lower in the latter. Clusterin and S-protein co-localized with MAC, respectively, in 20 and 12 specimens, and were not found in the absence of MAC. In addition S-protein deposits were seen only in biopsies positive for clusterin. Deposits of clusterin and S-protein did not correlate with the presence or absence of lesions. After irradiation with PTDL, the immediate complement activation was accompanied by MAC deposits that were granular and clearly located on vascular endothelial cells. Clusterin and S-protein were not present on these cells. In summary, clusterin localizes with MAC along the skin dermal-epidermal junction in patients with a positive LBT, suggesting that it has a similar and possibly more important role than S-protein in regulating immune complex-mediated MAC formation. By contrast, clusterin and S-protein are not involved at the time of MAC formation in cells undergoing necrosis after PTDL therapy.

Complement activation by pulsed tunable dye laser in normal skin and hemangioma.

Pulsed tunable dye laser (577 nm) (PTDL) therapy induces hemoglobin coagulation and tissue necrosis, which is mainly limited to blood vessels. To define whether this treatment activates complement in normal skin and senile hemangioma, we analyzed complement deposition in blood vessels by immunofluorescence. C3 fragments, C8, and C9 were detected with specific polyclonal antibodies. The membrane attack complex of complement (MAC) was demonstrated with a monoclonal antibody which reacts only with a neoantigen of MAC. Amplification of C3 deposition by the alternative pathway was determined on cryostat sections by indirect immunofluorescence with use of C4 deficient guinea pig (GP) serum. Normal skin and hemangiomas from three individuals were studied. In PTLD-irradiated normal skin, the main findings were as follows: 1) C3 fragments, C8, C9, and MAC were deposited in vessel walls; 2) these deposits were not due to denaturation of the proteins since they became apparent only 7 min after irradiation, contrary to immediate deposition of transferrin at the sites of erythrocyte coagulates; 3) the C3 deposits were shown to amplify complement activation by the alternative pathway, a reaction which was specific since tissue necrosis itself did not lead to such amplification; 4) these reactions preceded the local accumulation of polymorphonuclear leucocytes. Tissue necrosis was more pronounced in the hemangiomas. The larger angiomatous vessels in the center of the necrosis did not fix complement significantly. By contrast, complement deposition in the vessels situated at the periphery was similar to that observed in normal skin with one exception: C8, C9, and MAC were detected in some blood vessels immediately after laser treatment, a finding consistent with assembly of the MAC occurring directly without the formation of a C5 convertase. These results indicate that complement is activated in PTDL-induced vascular necrosis, and might be responsible for the ensuing inflammatory response.

Visible action spectrum for melanin-specific selective photothermolysis.

The skin of black and albino guinea pigs was irradiated with single, 750 nsec-long laser pulses at 435, 488, 530, and 560 nm in order to determine an action spectrum for the gross threshold response of immediate epidermal whitening. In addition, the immediate and delayed gross and histologic changes induced at, above, and below the threshold radiant exposures at all four wavelengths were studied. The action spectrum in the black guinea pigs was consistent with the reported absorption spectrum of DOPA-melanin. Histologically, there was epidermal damage immediately after radiant exposures at and above threshold at all four wavelengths. In addition, radiant exposures greater than threshold caused an immediate decrease in stainable epidermal pigment that was most marked at 435 and 488 nm. The healing response was also wavelength- and dose-dependent. Seven days after above-threshold exposures, there was little epidermal pigment in the 435 nm specimens. As wavelength increased, there was progressively more pigment, and in the 560 nm specimens, the epidermal pigment was equivalent to that seen in nonirradiated black guinea pig control specimens. Seven days after subthreshold radiant exposures, there was increased epidermal pigmentation and melanocytes at all four wavelengths. This was the most pronounced in the 435 nm specimens. There was no observable epidermal damage in albino guinea pig skin.

Pigmented guinea pig skin irradiated with Q-switched ruby laser pulses. Morphologic and histologic findings.

Q-switched ruby laser pulses cause selective damage to cutaneous pigmented cells. Repair of this selective damage has not been well described. Therefore, using epilated pigmented and albino guinea pig skin, we studied the acute injury and tissue repair caused by 40-ns, Q-switched ruby laser pulses. Gross observation and light and electron microscopy were performed. No specific changes were evident in the albino guinea pigs. In pigmented animals, with radiant exposures of 0.4 J/cm2 or greater, white spots confined to the 2.5-mm exposure sites developed immediately and faded over 20 minutes. Delayed depigmentation occurred at seven to ten days, followed by full repigmentation by four to eight weeks. Regrowing hairs in sites irradiated at and above 0.4 J/cm2 remained white for at least four months. Histologically, vacuolation of pigment-laden cells was seen immediately in the epidermis and the follicular epithelium at exposures of 0.3 J/cm2 and greater. Melanosomal disruption was seen immediately by electron microscopy at and above 0.3 J/cm2. Over the next seven days, epidermal necrosis was followed by regeneration of a depigmented epidermis. By four months, melanosomes and melanin pigmentation had returned; however, hair follicles remained depigmented and devoid of melanocytes. This study demonstrates that selective melanosomal disruption caused by Q-switched ruby laser pulses leads to transient cutaneous depigmentation and persistent follicular depigmentation. Potential exists for selective treatment of pigmented epidermal and dermal lesions with this modality.

The treatment of port-wine stains by the pulsed dye laser. Analysis of pulse duration and long-term therapy.

A flashlamp-pumped pulsed dye laser at 577 nm was evaluated in the treatment of port-wine stains. The degree of lesional lightening was compared following laser exposure with pulse durations of 20 and 360 microseconds. In addition, lesional therapy using the 360-microseconds pulse duration was evaluated for lightening and side effects following long-term patient observation and after repeated treatments of the same site. A total of 52 patients with port-wine stain were treated; their average age was 29 years, with eight patients less than 18 years, of whom 29 had comparative test site placement for the different pulse durations. Of these 29 patients, 25 demonstrated greater lightening at the 360-microseconds pulse duration test site. All 52 patients proceeded to receive full treatment placement with the 360-microseconds pulse duration, which resulted in an overall lightening of 42% after the initial treatment and 68% after re-treatment sessions. Forty-four percent of the patients had equal to or greater than 75% lesional lightening. Pretreatment anesthesia was unnecessary and only minimum posttreatment care was required. Mild adverse effects of epidermal change, depression, or pigmentary change appeared in only four cases and was limited to less than a 2% area in each of these lesions. These side effects did not recur when the lesions were re-treated at lower energy dosages. No posttreatment sclerosis or scarring appeared, even after multiple retreatment sessions to the same area, regardless of the anatomic location, color of the lesion, or age of the patient.(ABSTRACT TRUNCATED AT 250 WORDS)

Melanosomes are a primary target of Q-switched ruby laser irradiation in guinea pig skin.

The specific targeting of melanosomes may allow for laser therapy of pigmented cutaneous lesions. The mechanism of selective destruction of pigmented cells by various lasers, however, has not been fully clarified. Black, brown, and albino guinea pigs were exposed to optical pulses at various radiant exposure doses from a Q-switched, 40 nsec, 694 nm ruby laser. Biopsies were analyzed by light and electron microscopy (EM). Albino animals failed to develop clinical or microscopic evidence of cutaneous injury after irradiation. In both black and brown animals, the clinical threshold for gross change was 0.4 J/cm2, which produced an ash-white spot. By light microscopy, alterations appeared at 0.3 J/cm2 and included separation at the dermoepidermal junction, and the formation of vacuolated epidermal cells with a peripheral cytoplasmic condensation of pigment. By EM, enlarged melanosomes with a central lucent zone were observed within affected epidermal cells at 0.3 J/cm2. At 0.8 and 1.2 J/cm2, individual melanosomes were more intensely damaged and disruption of melanosomes deep in the hair papillae was observed. Dermal-epidermal blisters were formed precisely at the lamina lucida, leaving basal cell membranes and hemidesmosomes intact. Possible mechanisms for melanosomal injury are discussed. These observations show that the effects of the Q-switched ruby laser are melanin-specific and melanin-dependent, and may be useful in the selective destruction of pigmented as well as superficial cutaneous lesions.

[Selective photothermolysis: contribution to the treatment of flat angiomas (port wine stains) by laser]. / Photothermolyse sélective: application au traitement des angiomes plans par le laser.

Since 1962, lasers have been used in dermatology and have become the first choice in the treatment of superficial, vascular ectasia. Lasers are unique sources of light; they are coherent, monochromatic, collimated and intense. By careful selection of wavelength, pulse duration, and intensity, it is often possible to selectively confine a laser effect to a specific histologic structure in tissue, depending upon the tissue properties. The ideal treatment of Port Wine Stains (PWS) should irreversibly damage the ectatic vessels but minimize heating of the epidermis and superficial dermis. A theory, called selective photothermolysis, predicts the optimal combination of laser parameters of achieving this ideal treatment of PWS to be a wavelength of 577 nm, a pulse duration of 0.35-10 msec, and an energy per surface area of about 7-8 J/cm2. Laser wavelength: The wavelength of 577 nm is preferred because it: maximizes the selective absorption by hemoglobin, minimizes absorption by epidermal melanin, provides sufficient depth of penetration in the blood to coagulate 0.1 mm vessels allows penetration of light into dermis up to 1 mm. Laser pulse duration: A pulse-width in the range of 0.35-10 msec allows the temperature elevation to be uniform inside the vessel and to be confined to the vessel area. Shorter pulses superheat the red blood cells causing explosive boiling and hemorrhage. Longer pulses allow heat to diffuse away from vessels, requiring greater energies per pulse to achieve vessel damage. An increased energy per pulse increases the risk of excessive damage to surrounding tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Tunable pulsed dye laser for the treatment of benign cutaneous vascular ectasia.

A tunable pulsed dye laser emitting at 577 nm and 360 microseconds pulse width was used to treat benign cutaneous vascular ectasias other than port wine stain in 77 patients. Except for leg telangiectasias (34 patients), the overall response was excellent. Forty-two of forty-five patients with hemangiomas, spider nevi, angioma serpiginosum, venous lakes or facial telangiectasias showed excellent results after 1-4 consecutive treatments. Scarring was observed in none of the patients. These results confirm previous data on the use of the tunable dye laser in the treatment of port wine stain, and suggest that 577 nm wavelength and 360 microseconds pulse width allow the selective photothermolysis of vascular cutaneous ectasias with better clinical results than previously reported.