Establishment and Morphological Characterization of Patient-Derived Organoids from Breast Cancer.

BACKGROUND: Patient-derived organoids (PDO) technology represents an emerging tool for the study of tumor biology and drug responsiveness, thus being useful to design personalized medicine approaches. Despite several studies and clinical trials are ongoing using PDO from colorectal and pancreatic cancer, only few research papers have been published exploiting PDO from breast cancer. Here, we have developed a new protocol to establish PDO from surgical and biopsy samples. Furthermore, we have set up also the methodologies adopted for culture and morphological evaluations. RESULTS: Surgical and core biopsy specimens collected from 33 patients with diagnosis of breast cancer have been processed using the protocols here described obtaining PDO from cancerous and healthy mammary tissue (when available) in a quick and easy way with good yields. The more critical aspects influencing the yield were the characteristic of the tissue of origin (healthy vs tumor tissue) and the amount of material obtained after enzymatic digestion process. Success rate from healthy samples was about 20,83%, while this percentage was higher in samples from cancer tissue (i.e. 87,5%). Also the morphological characterization of breast cancer PDO by brightfield and transmission electron microscopy has been reported. CONCLUSIONS: Despite obtaining some organoids from a surgical or biopsy specimen is not a difficult procedure, the establishment of a stable organoid line able to grow and replicate, suitable for long-term biobank storage, is not so obvious. A novel, simple and quick procedure to obtain PDO from surgical and biopsy samples is here proposed to achieve high success rate .

Dermoscopy vs. reflectance confocal microscopy for the diagnosis of lentigo maligna.

BACKGROUND: Several dermoscopic and in vivo reflectance confocal microscopy (RCM) diagnostic criteria of lentigo maligna (LM)/lentigo maligna melanoma (LMM) have been identified. However, no study compared the diagnostic accuracy of these techniques. OBJECTIVE: We evaluated the diagnostic accuracy of dermoscopy and RCM for LM/LMM using a holistic assessment of the images. METHODS: A total of 223 facial lesions were evaluated by 21 experts. Diagnostic accuracy of the clinical, dermoscopic and RCM examination was compared. Interinvestigator variability and confidence level in the diagnosis were also evaluated. RESULTS: Overall diagnostic accuracy of the two imaging techniques was good (area under the curve of the sROC function: 0.89). RCM was more sensitive (80%, vs. 61%) and less specific (81% vs. 92%) than dermoscopy for LM/LMM. In particular, RCM showed a higher sensitivity for hypomelanotic and recurrent LM/LMM. RCM had a higher interinvestigator agreement and a higher confidence level in the diagnosis than dermoscopy. CONCLUSION: Reflectance confocal microscopy and dermoscopy are both useful techniques for the diagnosis of facial lesions and in particular LM/LMM. RCM is particularly suitable for the identification of hypomelanotic and recurrent LM/LMM.

Reflectance confocal microscopy analysis of equivocal melanocytic lesions with severe regression.

BACKGROUND: The differential diagnosis between regressing nevi and melanoma might be challenging; regressing areas can represent a confounding factor for the diagnosis and the histology still remain mandatory to rule out melanoma. Reflectance confocal microscopy may add valuable information by revealing features suggestive of the nature of the melanocytic proliferation. OBJECTIVE: To assess the impact of confocal microscopy in the management of regressive melanocytic lesions. METHODS: The dermoscopic analysis of 92 melanocytic lesions showing that more than 30% of regressions have been retrospectively considered, among them, 32 melanocytic lesions with a 7 check point list ≥3 they were assessed at the rcm and subsequently excised. For each selected lesion, dermoscopic features of regression (white scar-like areas, blue areas, blue white areas), distribution of regressing areas (central, peripheral, or both) and the percentage of regression have been examined by an expert in dermoscopy, blinded to the histological and confocal diagnosis. Subsequently, two experts in confocal microscopy revaluated, blinded from histology, RCM images. RESULTS: Of the 32 lesions analyzed, 23 (71.5%) were diagnosed histologically as nevi, and 9 (28.5%) as melanomas. 26 of 32 lesions (81.5%) exhibited regression >50% of the overall. On RCM, 11 lesions have been interpreted as malignant and 21 as benign. On RCM the majority of nevi exhibited regular architecture without cytological atypia. Epidermal disarray, pagetoid infiltration, disarranged dermo-epidermal junction architecture and atypical nests were considered as suspicious for malignancy. Good concordance between confocal readers has been detected. CONCLUSION: A combined dermoscopic/confocal approach can be used for the management of lesions exhibiting dermoscopic features of regression in order to provide a more conclusive pre-histological diagnosis avoiding a high number of unnecessary excisions. Limits of this study were represented by the relatively small number of lesions and the retrospective approach. Further, prospective studies on a larger number of cases, will be necessary in order to compare the efficacy of dermoscopy alone versus dermoscopy in combination with RCM for the evaluation of regression, suspected pigmented lesions.

Reflectance Confocal Microscopy for Inflammatory Skin Diseases.

In vivo reflectance confocal microscopy (RCM) is a relatively novel non-invasive tool for microscopic evaluation of the skin used prevalently for diagnosis and management of skin tumour. Its axial resolution, its non-invasive and easy clinical application represents the goals for a large diffusion of this technique. During the last 15 years, RCM has been demonstrated to be able to increase the sensibility and sensitivity of dermoscopy in the diagnosis of skin tumours integrating in real time clinic, dermoscopic and microscopic information useful for the definition of malignancy. Despite to date, no large comparative studies on inflammatory skin diseases has been published in the literature, several papers already showed that RCM has a potential for the evaluation of the descriptive features of the most common inflammatory skin diseases as psoriasis, lupus erythematosus, contact dermatitis and others. The aim of the application of this technique in non-neoplastic skin diseases has been prevalently focused on the possibility of clinical diagnosis confirmation, as well as therapeutic management. Moreover, the use of RCM as driver for an optimised skin biopsy has been also followed in order to reduce the number of unsuccessful histopathological examination. In this review article we describe the confocal features of the major groups of inflammatory skin disorders focusing on psoriasiform dermatitis, interface dermatitis and spongiotic dermatitis.

Reflectance confocal microscopy for inflammatory skin diseases.

Reflectance confocal microscopy evaluation of inflammatory skin diseases represents a relatively new indication that, during the last 5 years, has shown an increasing interest with consequent progressive increment of publications in literature. The success of RCM in this filed of dermatology is directly related to the high needing of non-invasive techniques able to reduce the number of skin biopsies and support the clinical diagnosis and patient's management. RCM demonstrated to visualize microscopic descriptors of inflammatory and pigmentary skin conditions with good reproducibility between observer and high grade of correspondence with optical histology. Moreover, RCM has shown to provide sufficient data to support clinical diagnosis and differential diagnosis of inflammatory and pigmentary skin diseases. Recently, several works published in literature have opened the prospective to use RCM also for therapeutic follow-up in order to monitor the improvement of the microscopic parameters and help to prevent treatment side effects. In this review article we present some examples of RCM application in inflammatory and pigmentary diseases.

Reflectance confocal microscopy for plaque psoriasis therapeutic follow-up during an anti-TNF-α monoclonal antibody: an observational multicenter study.

BACKGROUND: Psoriasis is a chronic recurrent inflammatory skin disease that affects 2-3% of the world population. Biologics are relatively new systemic treatments that block molecular steps important in the pathogenesis of psoriasis. In vivo Reflectance confocal microscopy (RCM) is a non-invasive, imaging technique already reported to be useful in the evaluation of the follow-up of PP under treatment with topical actives and phototherapy. No reports on systemic treatments have been proposed in literature so far. OBJECTIVE: The aim of this study was to evaluate the feasibility of RCM in the monitoring of microscopic response to a subcutaneous anti-TNF treatment, Adalimumab. METHODS: One target lesion with typical clinical aspect, from 48 psoriatic patients, was evaluated using RCM at baseline, after 4 and 8 weeks of treatment. RESULTS: Microscopic confocal changes were followed up during treatment. Results disclosed identification of early microscopic evidence of the anti-inflammatory activity of Adalimumab not detected at clinical examination. Confocal feature related to the effect of TNF-α on melanocytes activity has been also identified. CONCLUSION: Early detected RCM parameters related to Adalimumab activity could be used to identify an early response to the treatment. RCM seems to be able to give useful and practical information about follow-up in patients with PP under treatment with Adalimumab.

Randomized, double-blinded, vehicle-controlled, split-face study to evaluate the effects of topical application of a Gold Silk Sericin/Niacinamide/Signaline complex on biophysical parameters related to skin ageing.

OBJECTIVE: To investigate the effects of topical application of a Gold Silk Sericin (GSS) complex on biophysical parameters related to skin ageing. METHODS: A range of non-invasive bioengineering methods were deployed in an 8-week randomized, double-blinded, vehicle-controlled, split-face study among 40 female subjects aged 40-70. Endpoints measured included expert grades of skin condition, stratum corneum (SC) hydration, SC barrier function, elasticity and surface topography. RESULTS: The GSS complex produced significant single-variable (P < 0.05) improvements in SC hydration, barrier function, elasticity and surface topography compared with the Vehicle control. CONCLUSION: The GSS complex examined in this study represents an interesting new cosmetic topical technology with which to address multiple aspects of aged/photoaged female facial skin.

Real-time, non-invasive microscopic confirmation of clinical diagnosis of bullous pemphigoid using in vivo reflectance confocal microscopy.

BACKGROUND: Bullous pemphigoid is an autoimmune disease affecting prevalently the elder. In vivo reflectance confocal microscopy is a non-invasive technique for real-time imaging of the skin with cellular-level resolution. No previous data has been reported about confocal microscopy of bullous pemphigoid. Aim of this preliminary study is the evaluation of the potential of in vivo reflectance confocal microscopy for real-time, microscopical confirmation of clinical bullous pemphigoid diagnosis. METHODS: A total of nine lesions from patients affected by pemphigoid underwent in vivo reflectance confocal microscopy before histological examination. RESULTS: In our preliminary study, confocal microscopy showed high grade of correspondence to histopathology. In particular, presence of sub-epidermal cleft and variable amount of oedema of the upper dermis associated with inflammatory cells infiltration were seen as prevalent confocal features in the bullous lesions considered. Differently, in urticarial lesions, no specific features could be appreciated at confocal analysis beside the presence of signs of spongiosis and perivascular inflammation. CONCLUSION: Confocal microscopy seems to be useful for in vivo, microscopical confirmation of the clinical suspect of bullous pemphigoid and for biopsy site selection in urticarial lesions to obtain a more significant specimen for histopathological examination.