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    "textoCompleto" => "<span class="elsevierStyleSections"><p id="par0005" class="elsevierStylePara elsevierViewall">Melasma is a frequent acquired chronic dyschromia that affects photoexposed areas&#44; especially in women of childbearing age&#46; Ultraviolet radiation &#40;UVR&#41; is the main environmental stimulus that induces melasma pigmentation&#59; however&#44; the role of visible light &#40;VL&#58; 400&#8211;700&#8239;nm&#41; in its pathogenesis remains uncertain&#46; VL comprises about 44&#37; of solar radiation and promotes longer-lasting pigmentation &#40;&#62; 2 weeks&#41; than UVA&#59; however&#44; only in higher phototypes &#40;IV to VI&#41;&#46;<a class="elsevierStyleCrossRef" href="#bib0005"><span class="elsevierStyleSup">1</span></a></p><p id="par0010" class="elsevierStylePara elsevierViewall">Opsin-3&#44; also known as encephalopsin or panopsin &#40;OPN3&#41;&#44; is a G-protein-coupled photoreceptor which promotes blue light-induced melanogenesis &#40;420&#8211;490&#8239;nm&#41;&#46; Opsins 1 to 5 have been described in the retina&#59; however&#44; OPN3 is the most often expressed form in the skin&#44; which induces the phosphorylation of the microphthalmia-associated transcription factor &#40;MITF&#41; in melanocytes&#44; resulting in tyrosinase activation&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a> To date&#44; the differential expression of OPN3 in melasma skin compared to adjacent skin has not been investigated&#46;</p><p id="par0015" class="elsevierStylePara elsevierViewall">In the present study&#44; after the Ethics Committee approval&#44; 20 women with facial melasma &#40;diagnosed clinically by an experienced dermatologist&#41;&#44; without treatment for more than 30 days &#40;except sunscreen&#41; were submitted to two biopsies &#40;3&#8239;mm&#41;&#58; skin with melasma and unaffected adjacent skin &#40;&#60; 2&#8239;cm of distance&#41;&#46;</p><p id="par0020" class="elsevierStylePara elsevierViewall">The 40 specimens were submitted to triple-labeling immunofluorescence using primary antibodies and standardized dilutions&#58; rabbit anti-OPN3 antibody &#40;ab228748-Abcam&#44; Cambridge-MA&#44; USA&#41; 1&#58; 150&#59; mouse anti-vimentin antibody &#40;ab8978-Abcam&#44; Cambridge-MA&#44; USA&#41; 1&#58; 100&#44; which is expressed in melanocytes and fibroblast intermediate filaments&#59; and DAPI &#40;4&#39;&#44;6-Diamidino-2-Phenylindole&#41;&#44; for cell nucleus labeling&#46; Secondary antibodies &#40;chromophores&#41; and dilutions for OPN3&#58; AlexaFluor 594 - 1&#58; 500 &#40;red&#41;&#59; and&#44; for vimentin&#58; AlexaFluor 488 - 1&#58; 500 &#40;green&#41;&#46; All reactions were tested against positive and negative controls&#46;</p><p id="par0025" class="elsevierStylePara elsevierViewall">Three areas showing greater immunolabeling in the microscopic slides in each topography &#40;melasma and unaffected skin&#41; were photographed using LEICA TCS-SP8 laser confocal microscopy&#46; Using the red laser channel &#40;OPN3 chromophore&#41;&#44; guided by the colocalization of cytoplasmic green dots&#44; and outside the nuclear blue colocalization&#44; the average intensities of the image histograms &#40;range 0 to 255&#41; were estimated using the ImageJ software for the different cell types&#44; blinded to the topography&#46; The mean fluorescence intensity in each cell group was compared between the topographies by Student&#39;s <span class="elsevierStyleItalic">t</span>-test &#40;dependent data&#41;&#46; Cell counts between topographies were also estimated&#46; A two-tailed p-value &#60; 0&#46;05 was considered significant&#46;</p><p id="par0030" class="elsevierStylePara elsevierViewall">OPN3 receptors were identified in all studied cells&#58; keratinocytes&#44; basal layer melanocytes&#44; pendulous melanocytes&#44; and upper dermis fibroblasts &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>&#41;&#46; There was no differential expression in any cell group when comparing skin with melasma with the healthy adjacent skin &#40;<a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a>&#41;&#46;</p><elsevierMultimedia ident="fig0005"></elsevierMultimedia><elsevierMultimedia ident="tbl0005"></elsevierMultimedia><p id="par0035" class="elsevierStylePara elsevierViewall">There was no difference in melanocyte and fibroblast count in the upper dermis using a high magnification field &#40;&#215;400&#41; between the two topographies&#46; However&#44; the skin with melasma showed a higher density of pendulous melanocytes &#40;<a class="elsevierStyleCrossRef" href="#tbl0010">Table 2</a>&#41;&#46;</p><elsevierMultimedia ident="tbl0010"></elsevierMultimedia><p id="par0040" class="elsevierStylePara elsevierViewall">The results of this study suggest that the expression of OPN3 does not justify the difference in skin pigmentation in melasma in comparison with the unaffected adjacent skin&#46; If the VL plays an important role in its pathogenesis&#44; it does not play it in relation to OPN3 expression&#46; In fact&#44; patients with facial melasma did not show any worsening of the lesions after exposure to the computer screen &#40;8&#8239;h&#47;day&#44; 5 consecutive days&#41;&#44; at a distance of 20&#8239;cm&#46;<a class="elsevierStyleCrossRef" href="#bib0015"><span class="elsevierStyleSup">3</span></a> Moreover&#44; the expression of opsins &#40;OPN 1 to 5 &#41; in the skin is known to differ according to the phototype&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a></p><p id="par0045" class="elsevierStylePara elsevierViewall">There is a synergistic effect between VL and UVR type A1 &#40;UVA1&#58; 340&#8211;400&#8239;nm&#41;&#46; When individuals with phototype IV to V are irradiated with isolated VL&#44; pigmentation occurs non-linearly with the irradiated dose&#46; When UVA1 is associated with VL&#44; there is more intense pigmentation and the response is directly proportional to the utilized dose&#46;<a class="elsevierStyleCrossRef" href="#bib0020"><span class="elsevierStyleSup">4</span></a> The synergism of VL and UVA1 in the differential pigmentation of melasma needs to be explored using specific experimental designs&#46; The occurrence of OPN3 in human tissues outside the retina is called &#8220;non-visual&#8221; expression&#44; whose functions are not yet fully understood&#46; In the skin&#44; in addition to the modulation of melanogenesis&#44; it promotes differentiation of keratinocytes and the activation of metalloproteinases &#40;MMP-1&#44; -2&#44; -3&#44; and -9&#41; by fibroblasts&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a> OPN3 can also be induced on the skin after blue light irradiation in scar tissue repair models&#44; suggesting that it plays a role in skin healing&#46;<a class="elsevierStyleCrossRef" href="#bib0025"><span class="elsevierStyleSup">5</span></a></p><p id="par0050" class="elsevierStylePara elsevierViewall">Pendulous melanocytes are basal layer melanocytes that protrude towards the upper dermis and are characteristic of skin with melasma&#44; although they are found to a lesser extent in the adjacent photodamaged skin&#46;<a class="elsevierStyleCrossRef" href="#bib0030"><span class="elsevierStyleSup">6</span></a> Their pathophysiological importance is not clear&#44; but their occurrence seems to be related to the increase in Metalloproteinase-2 and basement membrane fragmentation&#46; Their greater numbers in the skin with melasma reinforces the role of UVR in the pathophysiology of the disease&#44; since UVA promotes an inflammatory microenvironment in the upper dermis&#44; with increased metalloproteinase activity &#40;especially of MMP-2 and -9&#41; and basement membrane degradation&#44; which facilitates the protrusion of melanocytes towards the dermis&#46;<a class="elsevierStyleCrossRefs" href="#bib0035"><span class="elsevierStyleSup">7&#44;8</span></a></p><p id="par0055" class="elsevierStylePara elsevierViewall">This study has limitations related to the semi-quantitative <span class="elsevierStyleItalic">in situ</span> investigation technique&#44; which does not identify different isoforms &#40;splicing&#41; of the photoreceptors or detects functional alterations of its pathway&#46; There are other spindle cells in the upper dermis in addition to fibroblasts &#40;<span class="elsevierStyleItalic">e&#46;g</span>&#46;&#44; CD4 &#43; undifferentiated mesenchymal dendritic cells&#41; which&#44; although less numerous&#44; are not distinguishable by morphology or vimentin labeling&#46; Also&#44; the other opsins &#40;OPN-1&#44; -2&#44; -4&#44; and -5&#41; were not assessed&#44; although less expressed in normal skin&#46;</p><p id="par0060" class="elsevierStylePara elsevierViewall">In conclusion&#44; there is no differential expression of OPN3 in keratinocytes&#44; melanocytes&#44; or fibroblasts in facial melasma when compared to the unaffected adjacent skin&#46;</p><span id="sec0005" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0005">Financial support</span><p id="par0065" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleGrantSponsor" id="gs0005">FUNADERSP</span> &#8211; <span class="elsevierStyleItalic">Fundo de Apoio &#224; Dermatologia de S&#227;o Paulo</span>&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0010">Authors&#8217; contributions</span><p id="par0070" class="elsevierStylePara elsevierViewall">Ana Cl&#225;udia Cavalcante Esp&#243;sito&#58; Participation in the design and planning of the study&#59; collection&#44; analysis&#44; and interpretation of data&#59; drafting of the manuscript&#59; approval of the final version&#46;</p><p id="par0075" class="elsevierStylePara elsevierViewall">Nath&#225;lia Pereira de Souza&#58; Collection&#44; analysis&#44; and interpretation of data&#59; critical review of the manuscript&#59; approval of the final version&#46;</p><p id="par0080" class="elsevierStylePara elsevierViewall">Luciane Donida Bartoli Miot&#58; Design and planning of the study&#59; interpretation of data&#59; critical review of the manuscript&#59; approval of the final version&#46;</p><p id="par0085" class="elsevierStylePara elsevierViewall">H&#233;lio Amante Miot&#58; Participation in the design and planning of the study&#59; analysis and interpretation of data&#59; drafting of the manuscript&#59; approval of the final version&#46;</p></span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0015">Conflicts of interest</span><p id="par0090" class="elsevierStylePara elsevierViewall">None declared&#46;</p></span></span>"
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    "fechaRecibido" => "2020-04-10"
    "fechaAceptado" => "2020-05-15"
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      0 => array:2 [
        "etiqueta" => "&#9734;"
        "nota" => "<p class="elsevierStyleNotepara" id="npar0015">How to cite this article&#58; Esp&#243;sito ACC&#44; de Souza NP&#44; Miot LDB&#44; Miot HA&#46; Expression of OPN3 in fibroblasts&#44; melanocytes&#44; and keratinocytes of skin with facial melasma in comparison with unaffected adjacent skin&#46; An Bras Dermatol&#46; 2021&#59;96&#58;367&#8211;9&#46;</p>"
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        "etiqueta" => "&#9734;&#9734;"
        "nota" => "<p class="elsevierStyleNotepara" id="npar0020">Study conducted at the Departments of Dermatology and Pathology&#44; Faculdade de Medicina de Botucatu&#44; Universidade Estadual Paulista&#44; Botucatu&#44; SP&#44; Brazil&#46;</p>"
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Melasma&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">p-value<a class="elsevierStyleCrossRef" href="#tblfn0005"><span class="elsevierStyleSup">a</span></a>&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">Suprabasal epidermis&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">30&#46;4 &#40;10&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">33&#46;9 &#40;9&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#46;170&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
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                  \t\t\t\t">Basal layer melanocytes&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">23&#46;3 &#40;9&#46;6&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">26&#46;0 &#40;7&#46;9&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">0&#46;364&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
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                  \t\t\t\t">Pendulous melanocytes&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">23&#46;7 &#40;11&#46;8&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">22&#46;3 &#40;8&#46;8&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">Upper dermis fibroblasts&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">16&#46;6 &#40;10&#46;2&#41;&nbsp;\t\t\t\t\t\t\n
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          "en" => "<p id="spar0010" class="elsevierStyleSimplePara elsevierViewall">Evaluation of the mean expression &#40;standard deviation&#41; of OPN3 in the 20 participants&#44; according to the topography and cell types evaluated&#46;</p>"
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Unaffected skin&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-head\n
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">p-value<a class="elsevierStyleCrossRef" href="#tblfn0010"><span class="elsevierStyleSup">a</span></a>&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">Basal layer melanocytes&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">3&#46;2 &#40;1&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">3&#46;6 &#40;1&#46;5&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">0&#46;451&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">2&#46;3 &#40;0&#46;9&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t"><span class="elsevierStyleBold">0&#46;031</span>&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">Fibroblasts of the upper dermis&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">4&#46;5 &#40;2&#46;0&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">4&#46;4 &#40;1&#46;4&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">0&#46;915&nbsp;\t\t\t\t\t\t\n
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          "en" => "<p id="spar0015" class="elsevierStyleSimplePara elsevierViewall">Mean &#40;standard deviation&#41; cell counts &#40;melanocytes&#44; pendulous melanocytes&#44; and fibroblasts&#41; per high magnification field &#40;&#215;400&#41;&#46;</p>"
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    ]
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                    0 => array:2 [
                      "titulo" => "Visible light in photodermatology"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:4 [
                            0 => "S&#46; Narla"
                            1 => "I&#46; Kohli"
                            2 => "I&#46;H&#46; Hamzavi"
                            3 => "H&#46;W&#46; Lim"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:2 [
                      "doi" => "10.1039/c9pp00425d"
                      "Revista" => array:6 [
                        "tituloSerie" => "Photochem Photobiol Sci"
                        "fecha" => "2020"
                        "volumen" => "19"
                        "paginaInicial" => "99"
                        "paginaFinal" => "104"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/31922171"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
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              "etiqueta" => "2"
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                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Illuminating insights into opsin 3 function in the skin"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:3 [
                            0 => "L&#46;E&#46; Olinski"
                            1 => "E&#46;M&#46; Lin"
                            2 => "E&#46; Oancea"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:3 [
                        "tituloSerie" => "Adv Biol Regul"
                        "fecha" => "2020"
                        "volumen" => "75"
                      ]
                    ]
                  ]
                ]
              ]
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            2 => array:3 [
              "identificador" => "bib0015"
              "etiqueta" => "3"
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                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Short-term exposure to blue light emitted by electronic devices does not worsen melasma"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:5 [
                            0 => "L&#46; Duteil"
                            1 => "C&#46; Queille-Roussel"
                            2 => "J&#46;P&#46; Lacour"
                            3 => "H&#46; Montaudi&#233;"
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Research Letter
Expression of OPN3 in fibroblasts, melanocytes, and keratinocytes of skin with facial melasma in comparison with unaffected adjacent skin
Ana Cláudia Cavalcante Espósito
Autor para correspondência
anaclaudiaesposito@gmail.com

Corresponding author.
, Nathália Pereira de Souza, Luciane Donida Bartoli Miot, Hélio Amante Miot
Departments of Dermatology and Pathology, Faculty of Medicine, Universidade Estadual Paulista, Botucatu, SP, Brazil
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          "en" => "<p id="spar0005" class="elsevierStyleSimplePara elsevierViewall">Overlapping image of triple-labeling immunofluorescence on skin with melasma&#46; Labeling in green &#40;vimentin&#41;&#44; red &#40;Opsin-3&#41;&#44; blue &#40;cell nuclei&#41;&#46; White outline arrow&#58; melanocytes in the basal layer&#46; Full white arrow&#58; pendulous melanocytes&#46; Dashed ellipses&#58; upper dermis fibroblasts&#46;</p>"
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    "textoCompleto" => "<span class="elsevierStyleSections"><p id="par0005" class="elsevierStylePara elsevierViewall">Melasma is a frequent acquired chronic dyschromia that affects photoexposed areas&#44; especially in women of childbearing age&#46; Ultraviolet radiation &#40;UVR&#41; is the main environmental stimulus that induces melasma pigmentation&#59; however&#44; the role of visible light &#40;VL&#58; 400&#8211;700&#8239;nm&#41; in its pathogenesis remains uncertain&#46; VL comprises about 44&#37; of solar radiation and promotes longer-lasting pigmentation &#40;&#62; 2 weeks&#41; than UVA&#59; however&#44; only in higher phototypes &#40;IV to VI&#41;&#46;<a class="elsevierStyleCrossRef" href="#bib0005"><span class="elsevierStyleSup">1</span></a></p><p id="par0010" class="elsevierStylePara elsevierViewall">Opsin-3&#44; also known as encephalopsin or panopsin &#40;OPN3&#41;&#44; is a G-protein-coupled photoreceptor which promotes blue light-induced melanogenesis &#40;420&#8211;490&#8239;nm&#41;&#46; Opsins 1 to 5 have been described in the retina&#59; however&#44; OPN3 is the most often expressed form in the skin&#44; which induces the phosphorylation of the microphthalmia-associated transcription factor &#40;MITF&#41; in melanocytes&#44; resulting in tyrosinase activation&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a> To date&#44; the differential expression of OPN3 in melasma skin compared to adjacent skin has not been investigated&#46;</p><p id="par0015" class="elsevierStylePara elsevierViewall">In the present study&#44; after the Ethics Committee approval&#44; 20 women with facial melasma &#40;diagnosed clinically by an experienced dermatologist&#41;&#44; without treatment for more than 30 days &#40;except sunscreen&#41; were submitted to two biopsies &#40;3&#8239;mm&#41;&#58; skin with melasma and unaffected adjacent skin &#40;&#60; 2&#8239;cm of distance&#41;&#46;</p><p id="par0020" class="elsevierStylePara elsevierViewall">The 40 specimens were submitted to triple-labeling immunofluorescence using primary antibodies and standardized dilutions&#58; rabbit anti-OPN3 antibody &#40;ab228748-Abcam&#44; Cambridge-MA&#44; USA&#41; 1&#58; 150&#59; mouse anti-vimentin antibody &#40;ab8978-Abcam&#44; Cambridge-MA&#44; USA&#41; 1&#58; 100&#44; which is expressed in melanocytes and fibroblast intermediate filaments&#59; and DAPI &#40;4&#39;&#44;6-Diamidino-2-Phenylindole&#41;&#44; for cell nucleus labeling&#46; Secondary antibodies &#40;chromophores&#41; and dilutions for OPN3&#58; AlexaFluor 594 - 1&#58; 500 &#40;red&#41;&#59; and&#44; for vimentin&#58; AlexaFluor 488 - 1&#58; 500 &#40;green&#41;&#46; All reactions were tested against positive and negative controls&#46;</p><p id="par0025" class="elsevierStylePara elsevierViewall">Three areas showing greater immunolabeling in the microscopic slides in each topography &#40;melasma and unaffected skin&#41; were photographed using LEICA TCS-SP8 laser confocal microscopy&#46; Using the red laser channel &#40;OPN3 chromophore&#41;&#44; guided by the colocalization of cytoplasmic green dots&#44; and outside the nuclear blue colocalization&#44; the average intensities of the image histograms &#40;range 0 to 255&#41; were estimated using the ImageJ software for the different cell types&#44; blinded to the topography&#46; The mean fluorescence intensity in each cell group was compared between the topographies by Student&#39;s <span class="elsevierStyleItalic">t</span>-test &#40;dependent data&#41;&#46; Cell counts between topographies were also estimated&#46; A two-tailed p-value &#60; 0&#46;05 was considered significant&#46;</p><p id="par0030" class="elsevierStylePara elsevierViewall">OPN3 receptors were identified in all studied cells&#58; keratinocytes&#44; basal layer melanocytes&#44; pendulous melanocytes&#44; and upper dermis fibroblasts &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>&#41;&#46; There was no differential expression in any cell group when comparing skin with melasma with the healthy adjacent skin &#40;<a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a>&#41;&#46;</p><elsevierMultimedia ident="fig0005"></elsevierMultimedia><elsevierMultimedia ident="tbl0005"></elsevierMultimedia><p id="par0035" class="elsevierStylePara elsevierViewall">There was no difference in melanocyte and fibroblast count in the upper dermis using a high magnification field &#40;&#215;400&#41; between the two topographies&#46; However&#44; the skin with melasma showed a higher density of pendulous melanocytes &#40;<a class="elsevierStyleCrossRef" href="#tbl0010">Table 2</a>&#41;&#46;</p><elsevierMultimedia ident="tbl0010"></elsevierMultimedia><p id="par0040" class="elsevierStylePara elsevierViewall">The results of this study suggest that the expression of OPN3 does not justify the difference in skin pigmentation in melasma in comparison with the unaffected adjacent skin&#46; If the VL plays an important role in its pathogenesis&#44; it does not play it in relation to OPN3 expression&#46; In fact&#44; patients with facial melasma did not show any worsening of the lesions after exposure to the computer screen &#40;8&#8239;h&#47;day&#44; 5 consecutive days&#41;&#44; at a distance of 20&#8239;cm&#46;<a class="elsevierStyleCrossRef" href="#bib0015"><span class="elsevierStyleSup">3</span></a> Moreover&#44; the expression of opsins &#40;OPN 1 to 5 &#41; in the skin is known to differ according to the phototype&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a></p><p id="par0045" class="elsevierStylePara elsevierViewall">There is a synergistic effect between VL and UVR type A1 &#40;UVA1&#58; 340&#8211;400&#8239;nm&#41;&#46; When individuals with phototype IV to V are irradiated with isolated VL&#44; pigmentation occurs non-linearly with the irradiated dose&#46; When UVA1 is associated with VL&#44; there is more intense pigmentation and the response is directly proportional to the utilized dose&#46;<a class="elsevierStyleCrossRef" href="#bib0020"><span class="elsevierStyleSup">4</span></a> The synergism of VL and UVA1 in the differential pigmentation of melasma needs to be explored using specific experimental designs&#46; The occurrence of OPN3 in human tissues outside the retina is called &#8220;non-visual&#8221; expression&#44; whose functions are not yet fully understood&#46; In the skin&#44; in addition to the modulation of melanogenesis&#44; it promotes differentiation of keratinocytes and the activation of metalloproteinases &#40;MMP-1&#44; -2&#44; -3&#44; and -9&#41; by fibroblasts&#46;<a class="elsevierStyleCrossRef" href="#bib0010"><span class="elsevierStyleSup">2</span></a> OPN3 can also be induced on the skin after blue light irradiation in scar tissue repair models&#44; suggesting that it plays a role in skin healing&#46;<a class="elsevierStyleCrossRef" href="#bib0025"><span class="elsevierStyleSup">5</span></a></p><p id="par0050" class="elsevierStylePara elsevierViewall">Pendulous melanocytes are basal layer melanocytes that protrude towards the upper dermis and are characteristic of skin with melasma&#44; although they are found to a lesser extent in the adjacent photodamaged skin&#46;<a class="elsevierStyleCrossRef" href="#bib0030"><span class="elsevierStyleSup">6</span></a> Their pathophysiological importance is not clear&#44; but their occurrence seems to be related to the increase in Metalloproteinase-2 and basement membrane fragmentation&#46; Their greater numbers in the skin with melasma reinforces the role of UVR in the pathophysiology of the disease&#44; since UVA promotes an inflammatory microenvironment in the upper dermis&#44; with increased metalloproteinase activity &#40;especially of MMP-2 and -9&#41; and basement membrane degradation&#44; which facilitates the protrusion of melanocytes towards the dermis&#46;<a class="elsevierStyleCrossRefs" href="#bib0035"><span class="elsevierStyleSup">7&#44;8</span></a></p><p id="par0055" class="elsevierStylePara elsevierViewall">This study has limitations related to the semi-quantitative <span class="elsevierStyleItalic">in situ</span> investigation technique&#44; which does not identify different isoforms &#40;splicing&#41; of the photoreceptors or detects functional alterations of its pathway&#46; There are other spindle cells in the upper dermis in addition to fibroblasts &#40;<span class="elsevierStyleItalic">e&#46;g</span>&#46;&#44; CD4 &#43; undifferentiated mesenchymal dendritic cells&#41; which&#44; although less numerous&#44; are not distinguishable by morphology or vimentin labeling&#46; Also&#44; the other opsins &#40;OPN-1&#44; -2&#44; -4&#44; and -5&#41; were not assessed&#44; although less expressed in normal skin&#46;</p><p id="par0060" class="elsevierStylePara elsevierViewall">In conclusion&#44; there is no differential expression of OPN3 in keratinocytes&#44; melanocytes&#44; or fibroblasts in facial melasma when compared to the unaffected adjacent skin&#46;</p><span id="sec0005" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0005">Financial support</span><p id="par0065" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleGrantSponsor" id="gs0005">FUNADERSP</span> &#8211; <span class="elsevierStyleItalic">Fundo de Apoio &#224; Dermatologia de S&#227;o Paulo</span>&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0010">Authors&#8217; contributions</span><p id="par0070" class="elsevierStylePara elsevierViewall">Ana Cl&#225;udia Cavalcante Esp&#243;sito&#58; Participation in the design and planning of the study&#59; collection&#44; analysis&#44; and interpretation of data&#59; drafting of the manuscript&#59; approval of the final version&#46;</p><p id="par0075" class="elsevierStylePara elsevierViewall">Nath&#225;lia Pereira de Souza&#58; Collection&#44; analysis&#44; and interpretation of data&#59; critical review of the manuscript&#59; approval of the final version&#46;</p><p id="par0080" class="elsevierStylePara elsevierViewall">Luciane Donida Bartoli Miot&#58; Design and planning of the study&#59; interpretation of data&#59; critical review of the manuscript&#59; approval of the final version&#46;</p><p id="par0085" class="elsevierStylePara elsevierViewall">H&#233;lio Amante Miot&#58; Participation in the design and planning of the study&#59; analysis and interpretation of data&#59; drafting of the manuscript&#59; approval of the final version&#46;</p></span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0015">Conflicts of interest</span><p id="par0090" class="elsevierStylePara elsevierViewall">None declared&#46;</p></span></span>"
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        "nota" => "<p class="elsevierStyleNotepara" id="npar0015">How to cite this article&#58; Esp&#243;sito ACC&#44; de Souza NP&#44; Miot LDB&#44; Miot HA&#46; Expression of OPN3 in fibroblasts&#44; melanocytes&#44; and keratinocytes of skin with facial melasma in comparison with unaffected adjacent skin&#46; An Bras Dermatol&#46; 2021&#59;96&#58;367&#8211;9&#46;</p>"
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        "nota" => "<p class="elsevierStyleNotepara" id="npar0020">Study conducted at the Departments of Dermatology and Pathology&#44; Faculdade de Medicina de Botucatu&#44; Universidade Estadual Paulista&#44; Botucatu&#44; SP&#44; Brazil&#46;</p>"
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                  \t\t\t\t">30&#46;4 &#40;10&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">33&#46;9 &#40;9&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">0&#46;170&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">23&#46;3 &#40;9&#46;6&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">0&#46;364&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttop\n
                  \t\t\t\t">3&#46;2 &#40;1&#46;3&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">3&#46;6 &#40;1&#46;5&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#46;451&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">Pendulous melanocytes&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">2&#46;3 &#40;0&#46;9&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">1&#46;8 &#40;0&#46;7&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t"><span class="elsevierStyleBold">0&#46;031</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">Fibroblasts of the upper dermis&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">4&#46;5 &#40;2&#46;0&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">4&#46;4 &#40;1&#46;4&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#46;915&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr></tbody></table>
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          "en" => "<p id="spar0015" class="elsevierStyleSimplePara elsevierViewall">Mean &#40;standard deviation&#41; cell counts &#40;melanocytes&#44; pendulous melanocytes&#44; and fibroblasts&#41; per high magnification field &#40;&#215;400&#41;&#46;</p>"
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