The aim of this retrospective pilot study was to evaluate the Aperio nuclear V9 algorithm as an image analysis tool to observe the histopathological changes of ocular surface squamous neoplasia (OSSN). A histopathological assessment, including the Ki-67 proliferative index (PI) of immunohistochemically-stained tumor conjunctiva (TC) and healthy conjunctiva (HC) tissues, was performed in six cases of OSSN. The Aperio V9 algorithm was applied to digital images of the tissue specimens to count the Ki-67 PI and to measure the nuclear area indices. This digital algorithm was validated using stereological and visual analysis methods. The visual scoring of Ki-67 PI ranged from 22 to 60% (mean, 38.5%), and from 5 to 20% (mean 9.5%) in TC and HC tissue, respectively. The computer-aided analysis, using the Aperio nuclear V9 algorithm, revealed that the Ki-67 PI ranged from 21.5 to 43.5% (mean, 33.6%), and from 1.9 to 21.0% (mean, 11.8%) in the TC and HC tissue, respectively. The stereological method demonstrated that the Ki-67 PI ranged from 30.1 to 51.5% (mean, 41.0%), and from 3.2 to 30.1% (mean, 15.1%) in the TC and HC tissues, respectively. The strongest association in the collinearity of regression analysis was observed between the Aperio nuclear V9 algorithm/stereological models in the TC tissue (r2=0.7; P=0.04) and the HC tissue (r2=0.7; P=0.03), and the visual/stereological models in the TC tissue (r2=0.7; P=0.04) and the visual/Aperio nuclear V9 algorithm models in the HC tissue (r2=0.7; P=0.04). A weak and statistically insignificant association was identified between the visual/Aperio nuclear V9 algorithm analysis in the TC tissue (r2=0.4; P=0.2) and the visual/stereological models in the HC tissue (r2=0.5; P=0.13). No significant difference was observed between the nuclear area of the TC (mean, 36.5 μm2) and HC (mean, 35.7 μm2; P=0.88) tissues. It was concluded that the Aperio nuclear V9 algorithm is a useful tool for the reliable analysis of histopathological changes of OSSN. The results of this computer-aided algorithm correlate strongly with the stereological method when assessing the Ki-67 PI.

Purpose To investigate a tumor proliferation marker Ki‐67 in conjunctival malignancies and to compare the results of different quantitative assessments. Methods This study was designed as a pilot study to assess nuclear proliferation protein (Ki 67) in conjunctival malignancies. Immunohistochemical staining of Ki‐67 was evaluated in 8 surgical samples of ocular surface neoplasias: 6 cases of invasive conjunctival squamous carcinoma and 2 of conjuntival melanoma. Ki‐67 counting and evaluation was assessed on selected areas using different methods: manual counting (MC) of >1500 cells;digital image analysis with Nuclear V9 algorithm and digital image analysis using stereology module for ImageScope of Aperio. Ki‐67 values from manual, stereoscopic and automated analyses were compared. Results The MC, the nuclear V9 algorithm analysis and stereologic method revealed positivity of Ki‐67 in invasive conjunctival squamous carcinoma from 22 to 60%, 21.5 to 43.5 % and from 30.1 to 51.5% respectively; in conjunctival melanomas from 20 to 30 %; from 17.8 to 18%, and from 7.17 to 29.1% respectively. The strongest correlation was observed between Aperio algorithm and stereologic method in conjunctival squamous carcinoma (0.87). Weaker correlation was found between Aperio/manual (0.6) and manual/stereological (0.83) methods. Conclusion Our data demonstrate that different methods of Ki‐67 evaluation may give dissimilar results, but results of this computer assisted nuclear V9 algorithm correlates strongly with stereoscopic analysis and can be helpful in accurate quantitative evaluation of imunochistochemically stained biomarkers.

Purpose To analyze and evaluate biological and morphometrical changes and differences of healthy and carcinomatous conjunctiva using Aperio nuclear v9 algorithm. Methods A pilot study for the objective evaluation of carcinomatous conjunctiva in 6 cases. There were analyzed 6 excised conjunctival tumors specimens from 6 patients (6 eyes). Medium age of the patients was 68 years. There were two males and four females. In 3 patients (3 eyes) conjunctival tumor was excised, cryoaplication and amniotic membrane transplantation was performed. In 3 patients excisional biopsy was performed without additional surgical manipulations. Conjunctival specimens were stained with hematoxilin and eosin. Immunohistochemical (IHC) staining with a proliferation marker of cells Ki‐67 was performed for all specimens. Digital slides of the IHC of Ki‐67 from these biopsies were obtained by Aperio ScanScope GL. Aperio Nuclear V9 algorithm was applied to assay proliferation indices (PI) of nuclei stained with Ki‐67 and size of all nuclei in both abnormal and normal conjunctiva. Results There were analyzed from 1585 till 5020 malignant and 242‐714 normal cells. PI of Ki‐67 ≥19% (range 19.3‐62.1%) was found in carcinomatous conjunctiva comparing with normal conjunctiva PI ≤13.5% (range 1.9‐21.6%) in the same patients. No significant difference was found between nucleus size in pathological (mean 41 µm2, range 33‐64 µm2) and normal conjunctiva (mean 35.5 µm2, range 34‐36 µm2). Conclusion Aperio nuclear algorithm can be used for carcinomatous and healthy conjunctiva analysis. Further studies of the malignant and healthy eye tissues using larger cohort are required.

Purpose To report two cases of Purtscher‐like retinopathy associated with acute pancreatitis. Methods Retrospective analysis of two cases of Purtscher‐like retinopathy associated with acute pancreatitis. Fluorescein angiography (FAG), retinal photography, laboratory tests, computer tomography (CT) were analyzed. Results 2 cases were identified in a period from 2007 to 2013. The first, 57 y.o patient with a history of recently treated acute pancreatitis, was referred to clinic 2 weeks after visual impairment. Visual acuity (VA) was 0,3 in the right eye and 0,25 in the left eye. FAG revealed prolonged arterio‐venous passage time, enlarged foveal avascular zone (FAZ), fluorescein blockage temporal to optic nerve disc due to soft exudates. Patient was treated with systemic steroids intravenously. VA improved to 1,0 after 9 days of treatment. The second, 39 years old patient was referred to our clinic 3 days after visual impairment. At the time of hospitalization despite patients normal vital signs, laboratory tests and CT revealed acute pancreatitis and reactive hepatitis. VA was hand motion in the right eye and 0,01 in the left eye. FAG showed large FAZ and evidence of leakage from vessels in the late phase. After treatment with intravenous steroids, VA improved to counting fingers at 30 cm in the right eye and 0,03 in the left eye. Conclusion Prognosis of Purtscher‐like retinopathy may depend on systemic disease severity, VA in early stages and the presence of macular ischemia. Pancreatitis‐associated retinopathy is rare and there are no evidence‐based therapeutic recommendations.