Syndromic and non-syndromic forms of retinitis pigmentosa: a comprehensive Italian clinical and molecular study reveals new mutations.

Mutations in more than 60 different genes have been associated with non-syndromic and syndromic retinitis pigmentosa (RP), a heterogeneous group of inherited retinal dystrophies. To increase the understanding of the molecular epidemiology of the disease in Italy, we analyzed 56 patients with syndromic and non-syndromic forms of RP attending the Retinitis Pigmentosa Center of San Paolo Hospital (Milan, Italy). Patients underwent detailed clinical examination. Genomic DNA isolated from peripheral blood samples was screened for mutations in different genes according to RP form by direct sequencing analysis. The impact of novel missense mutations on protein functions was predicted by in silico analysis and protein sequence alignment. Cosegregation analysis was performed between available family members. Forty-one of the 56 probands analyzed had non-syndromic and 15 had syndromic RP forms. Putative disease-causing mutations were identified in 19 of 56 unrelated RP probands. Mutation screening identified a total of 22 different heterozygous variants. Notably, 12 of these putative pathogenic mutations have not been previously reported. New variants were found to be located on the USH2A, RPGR, EYS, and RHO genes. All 3 new variants detected in X-linked RP probands were confirmed in other affected family members. We found a positivity rate of 24.4% and 60% for probands with non-syndromic and syndromic RP, respectively. This is the first report of RPGR X-linked RP proband-ORF15 mutations in Italian patients with X-linked (XL)-RP. In addition, this is the first report of data regarding the association between EYS mutations and non-syndromic RP forms in the Italian population.

Proliferative activity in human breast cancer: Ki-67 automated evaluation and the influence of different Ki-67 equivalent antibodies.

BACKGROUND: Ki67 labeling index (Ki67 LI), the percentage Ki67 immunoreactive cells, is a measure of tumor proliferation, with important clinical relevance in breast cancer, and it is extremely important to standardize its evaluation. AIM: To test the efficacy of computer assisted image analysis (CAIA) applied to completely digitized slides and to assess its feasibility in routine practice and compare the results obtained using two different Ki67 monoclonal antibodies. MATERIALS AND METHODS: 315 consecutive breast cancer routinely immunostained for Ki-67 (223 with SP6 and 92 with MM1 antibodies previously examined by an experienced pathologist, have been re-evaluated using Aperio Scanscope Xs. RESULTS: Mean human Ki67 LI values were 36%± 14.% and 28% ± 18% respectively for SP6 and MM1 antibodies; mean CAM Ki67 LI values were 31%± 19% and 22% ± 18% respectively for SP6 and MM1. Human and CAIA evaluation are statistically highly correlated (Pearson: 0.859, p<0.0001), although human LI are systematically higher. An interobserver variation study on CAIA performed on 84 cases showed that the correlation between the two evaluations was linear to an excellent degree. DISCUSSION: Our study shows that a) CAIA can be easily adopted in routine practice, b) human and CAIA Ki67 LI are highly correlated, although human LI are systematically higher, c) Ki67 LI using different evaluation methods and different antibodies shows important differences in cut-off values.