Elevated X-Box Binding Protein1 Splicing and Interleukin-17A Expression Are Associated With Active Generalized Vitiligo in Gujarat Population

Vitiligo is an autoimmune skin disorder defined by the destruction of functional epidermal melanocytes. It is a multifactorial and polygenic disorder caused due to oxidative stress, endoplasmic reticulum (ER) stress, and autoimmunity, among other factors. In the present study, we aimed to investigate the association of X-box Binding Protein 1 (XBP1) and Interleukin-17A (IL-17A) polymorphisms and monitor their systemic as well as skin expression levels in vitiligo patients from Gujarat population in India. XBP1 rs2269577 G/C, IL17A rs2275913 G/A and IL17A rs8193036 C/T polymorphisms were genotyped by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) method in 312 controls and 276 vitiligo patients. Transcript levels of spliced (sXBP1), unspliced XBP1 (uXBP1) and IL17A from peripheral blood mononuclear cells (PBMCs) as well as spliced and unspliced XBP1 from skin samples were analyzed by qPCR. IL-17A protein levels in suction-induced blister fluid (SBF) from the skin of study subjects were estimated by ELISA. The results revealed that genotype (p=0.010) and allele (p=0.014) frequencies of XBP1 rs2269577 G/C polymorphism were significantly different, however, no significant difference was observed in frequencies of IL17A rs2275913 G/A and IL17A rs8193036 C/T polymorphisms in control and patient population. Gene expression analysis revealed that sXBP1 and IL17A levels were significantly higher in PBMCs of generalized (p=0.030 and p=0.039, respectively) and active (p=0.024 and p=0.017, respectively) vitiligo patients. Moreover, we observed a significantly elevated sXBP1 expression (p=0.037) as well as IL-17A protein levels (p=0.009) in perilesional skin of vitiligo patients as compared to controls. Overall, these findings suggest XBP1 and IL17A play an important role in vitiligo and further substantiate the involvement of ER stress in exacerbating immune-mediated vitiligo pathogenesis.

Computer-Aided Detection (CADe) System with Optical Coherent Tomography for Melanin Morphology Quantification in Melasma Patients

Dark skin-type individuals have a greater tendency to have pigmentary disorders, among which melasma is especially refractory to treat and often recurs. Objective measurement of melanin amount helps evaluate the treatment response of pigmentary disorders. However, naked-eye evaluation is subjective to weariness and bias. We used a cellular resolution full-field optical coherence tomography (FF-OCT) to assess melanin features of melasma lesions and perilesional skin on the cheeks of eight Asian patients. A computer-aided detection (CADe) system is proposed to mark and quantify melanin. This system combines spatial compounding-based denoising convolutional neural networks (SC-DnCNN), and through image processing techniques, various types of melanin features, including area, distribution, intensity, and shape, can be extracted. Through evaluations of the image differences between the lesion and perilesional skin, a distribution-based feature of confetti melanin without layering, two distribution-based features of confetti melanin in stratum spinosum, and a distribution-based feature of grain melanin at the dermal–epidermal junction, statistically significant findings were achieved (p-values = 0.0402, 0.0032, 0.0312, and 0.0426, respectively). FF-OCT enables the real-time observation of melanin features, and the CADe system with SC-DnCNN was a precise and objective tool with which to interpret the area, distribution, intensity, and shape of melanin on FF-OCT images.

Serration pattern analysis as a practical adjunct tool for categorization of subepidermal autoimmune blistering diseases

Background: Serration pattern analysis helps in the classification of subepidermal autoimmune blistering disorders; more precisely, it helps to differentiate epidermolysis bullosa acquisita from other subepidermal autoimmune blistering disorders. Most of the published reports of this tool have come from a single center. Objectives: The objectives of the study were to study the utility of serration pattern analysis in classifying subepidermal autoimmune blistering disorders. Methods: Seventy five cases of subepidermal autoimmune blistering disorders were enrolled in this prospective study. A three millimeter punch biopsy was taken from the perilesional skin or mucosa for direct immunofluorescence; indirect immunofluorescence was carried out using salt-split skin. Subclassification of subepidermal autoimmune blistering disorders was done based on direct immunofluorescence, indirect immunofluorescence on salt-split skin, indirect immunofluorescence using knockout skin and serration pattern analysis findings. Results: Indirect immunofluorescence was positive in 68 cases; 14 cases showed a dermal staining pattern while the rest showed either an epidermal or a combined pattern. All patients with epidermal or combined staining patterns showed “n” serrated pattern on direct immunofluorescence. Nine patients with dermal staining on indirect immunofluorescence also revealed an “n” serration pattern on direct immunofluorescence indicating the diagnosis of anti-p200 pemphigoid, and the rest showed a “u” serrated pattern. Three patients with negative indirect immunofluorescence showed “u” serration on direct immunofluorescence while the rest showed “n” serration. Limitations: ELISA and immunoblotting could not be performed due to resource constraints. Conclusion: Based on indirect immunofluorescence and serration pattern analysis, classification of the majority of patients with subepidermal autoimmune blistering disorders was possible in our study. Pattern recognition is a cost-effective tool and can be easily learnt. It is recommended to be practiced in all laboratories where facilities for advanced immunological diagnosis are unavailable.

Establishment of a Three-Dimensional In Vitro Model of Equine Papillomavirus Type 2 Infection

There is growing evidence that equine papillomavirus type 2 (EcPV2) infection is etiologically associated with the development of genital squamous cell carcinoma (SCC) and precursor lesions in equids. However, the precise mechanisms underlying neoplastic progression remain unknown. To allow the study of EcPV2-induced carcinogenesis, we aimed to establish a primary equine cell culture model of EcPV2 infection. Three-dimensional (3D) raft cultures were generated from equine penile perilesional skin, plaques and SCCs. Using histological, molecular biological and immunohistochemical methods, rafts versus corresponding natural tissue sections were compared with regard to morphology, presence of EcPV2 DNA, presence and location of EcPV2 gene transcripts and expression of epithelial, mesenchymal and tumor/proliferation markers. Raft cultures from perilesional skin harboring only a few EcPV2-positive (EcPV2+) cells accurately recapitulated the differentiation process of normal skin, whilst rafts from EcPV2+ penile plaques were structurally organized but showed early hyperplasia. Rafts from EcPV2+ SCCs exhibited pronounced hyperplasia and marked dysplasia. Raft levels of EcPV2 oncogene transcription (E6/E7) and expression of tumor/proliferation markers p53, Ki67 and MCM7 expression positively correlated with neoplastic progression, again reflecting the natural situation. Three-dimensional raft cultures accurately reflected major features of corresponding ex vivo material, thus constituting a valuable new research model to study EcPV2-induced carcinogenesis.

Real-Time Confocal Imaging for Hidradenitis Suppurativa: Description of Morphological Aspects and Focus on the Role of Follicular Ostia

<b><i>Background:</i></b> Hidradenitis suppurativa (HS) is a chronic, recurrent, inflammatory skin disease involving the follicular unit. Growing evidence suggests a crucial role of hyperkeratosis, infundibular plugging and perifolliculitis in its pathogenesis. <b><i>Objectives:</i></b> To characterize the microscopic morphology of open comedos using in vivo reflectance confocal microscopy (RCM) and define alterations occurring in HS normal-appearing perilesional skin compared to healthy skin of a control group, discussing therefore microscopic differences. <b><i>Materials and Methods:</i></b> Twenty patients (11 males, 9 females; aged 19–51 years) affected by HS were recruited. RCM was performed on open comedos of axillary/inguinal regions and on normal-appearing skin areas distant at least 1.5 cm from HS lesions. Ten healthy individuals (6 males, 4 females) were included as control group. <b><i>Results:</i></b> RCM aspects of 78 open comedos were explored, observing circular/ovoidal structures disclosing a hyperrefractive amorphous material (72/78, 92.3%) within the infundibular cavity surrounded with a bright (51/78, 65.4%) and regular (52/78, 66.2%) border. Follicular ostia of HS perilesional skin (<i>n</i> = 541) compared to follicular ostia of a control group (<i>n</i> = 321) were characterized by a larger infundibular diameter (183.4 vs. 85.8 µm) and diverged in terms of material collected inside infundibula (44.5 vs. 21%) and keratinization of the border (47.6 vs. 25.5%). <b><i>Conclusion:</i></b> In vivo characterization of open comedos and the recognition of microscopic subclinical alterations of HS normal-appearing skin, compared to healthy skin, could add further insights into the definition of biological events leading to HS manifestations, including the dysregulated process of keratinization.

Aktinische Keratose: Drei therapeutische Verfahren im direkten Vergleich

<b>Background:</b> The efficacy for actinic keratosis (AK) clearance of field-directed treatments has been investigated in randomized studies against placebo, but the comparison of results is difficult for several methodological reasons. <b>Objectives:</b> The present study aims to compare efficacy of MAL-photodynamic therapy (MAL-PDT), ingenol mebutate gel (IMB) and diclofenac plus hyaluronate gel (DHA) on multiple AKs assessing a new performance index of efficacy, the cumulative AK area and evaluating dermoscopical and high-frequency ultrasound (HFUS) changes. <b>Methods:</b> Patients with ≥5 Olsen II AKs in a 25 cm2 area of the scalp and face were enrolled and randomized to one of the treatment choices. Number of AKs and cumulative area were assessed before and after treatment. Dermoscopy and HFUS were performed on a single AK and surrounding photo-damaged skin in the treatment area. <b>Results:</b> Cumulative AKs area reduced significantly more with PDT compared to other treatment options and with IMB in comparison to DHA. PDT was also the only treatment option that increased at a significant level the dermal density in both target AK and the surrounding skin and decreased significantly the SLEB thickness in the perilesional skin at HFUS. <b>Conclusions:</b> MAL-PDT is more effective than IMB and DHA for reducing the cumulative AK area which is calculated digitally from 3D pictures and should be the preferred performance index for the evaluation of the efficacy of treatments for AKs, rolling out clinical and dermoscopy evaluation. MAL-PDT improves all HFUS features of chronic photodamages of the dermis of the skin underlying and surrounding the AK spots.