SummaryThe aim of this investigation was to study the expression of caspases 3, 6 and 8 and their association to apoptosis in preneoplastic and neoplastic lesions of the breast. The material consisted of nine benign breast epithelial hyperplasias, 15 atypical hyperplasias, 74 in situ and 82 invasive carcinomas. The extent of apoptosis was assessed by the TUNEL method and caspase 3, 6 and 8 expression by immunohistochemistry with specific antibodies. Increased caspase 3 immunopositivity, as compared to staining of normal breast ductal epithelium, was seen in 22% of benign epithelial hyperplasias, 25% of atypical hyperplasias, 58% of in situ carcinomas and 90% of invasive carcinomas. The corresponding percentages for caspase 6 and 8 were 11%, 25%, 60%, 87% and 22%, 57%, 84%, 83% respectively. In highgrade in situ lesions there were significantly more cases with strong caspase 3, 6 and 8 immunoreactivity than in low-and intermediate-grade lesions (P = 0.0045, P = 0.049 and P = 0.0001 respectively). In invasive carcinomas, however, no association between a high tumour grade and caspase 3, 6 or 8 expression was found (P = 0.27, P = 0.26 and P = 0.69 respectively). The mean apoptotic index was 0.14 ± 0.14% in benign epithelial hyperplasias, 0.17 ± 0.12% in atypical hyperplasias, 0.61 ± 0.88% in in situ carcinomas and 0.94 ± 1.21% in invasive carcinomas. In all cases strong caspase 3, 6 and 8 positivity was significantly associated with the extent of apoptosis (P < 0.001, P = 0.015 and P = 0.050 respectively). The results show that synthesis of caspases 3, 6 and 8 is up-regulated in neoplastic breast epithelial cells in parallel to the increase in the apoptotic index and progression of the breast lesions.

Manganese superoxide dismutase (MnSOD) is an antioxidant enzyme capable of neutralizing superoxide anion molecules. In previous studies it has been suggested to suppress both tumour proliferation and apoptosis. This study investigated 65 invasive, 50 in situ and 19 benign hyperplastic breast lesions for its immunohistochemical expression. MnSOD expression was also tested with in situ hybridization. To study cell proliferation, apoptosis and their association with MnSOD expression the neoplastic breast lesions were immunostained with a monoclonal antibody to Ki-67 and the extent of apoptosis in them was determined by the TUNEL method. 32/65 (49%) of the invasive ductal carcinomas, 41/50 (82%) of the in situ and 15/19 (79%) of the benign hyperplasias expressed the MnSOD protein. There were significantly more MnSOD positive cases in in situ carcinoma and in benign hyperplasia than in invasive carcinoma (p=0.00016 and p=0.022, respectively). Positivity was also more frequently found in non-neoplastic ductal and acinar epithelial cells than in invasive carcinoma. On the other hand, neoplastic epithelial cells of invasive and in situ carcinoma showed strong positivity more often than the epithelial cells of benign hyperplasia or non-neoplastic epithelium. In breast lesions, MnSOD positivity did not associate with proliferation or apoptosis. The lower frequency of MnSOD positive cases in invasive breast carcinoma suggests that the lack of its expression might contribute to the development of an invasive breast carcinoma phenotype and that it could in this way operate as a tumour suppressor gene, as previously suggested.

Background: Neuroinflammation often develops in sepsis along with increasing permeability of the blood-brain barrier (BBB), which leads to septic encephalopathy. The barrier is formed by tight junction structures between the cerebral endothelial cells. We investigated the expression of tight junction proteins related to endothelial permeability in brain autopsy specimens in critically ill patients deceased with sepsis and analyzed the relationship of BBB damage with measures of systemic inflammation and systemic organ dysfunction. Methods: The case series included all (385) adult patients deceased due to sepsis in the years 2007-2015 with available brain specimens taken at autopsy. Specimens were categorized according to anatomical location (cerebrum, cerebellum). The immunohistochemical stainings were performed for occludin, ZO-1, and claudin. Patients were categorized as having BBB damage if there was no expression of occludin in the endothelium of cerebral microvessels. Results: Brain tissue samples were available in 47 autopsies, of which 38% (18/47) had no expression of occludin in the endothelium of cerebral microvessels, 34% (16/47) developed multiple organ failure before death, and 74.5% (35/47) had septic shock. The deceased with BBB damage had higher maximum SOFA scores (16 vs. 14, p = 0.04) and more often had procalcitonin levels above 10 μg/L (56% vs. 28%, p = 0.045) during their ICU stay. BBB damage in the cerebellum was more common in cases with C-reactive protein (CRP) above 100 mg/L as compared with CRP less than 100 (69% vs. 25%, p = 0.025). Conclusions: In fatal sepsis, damaged BBB defined as a loss of cerebral endothelial expression of occludin is related with severe organ dysfunction and systemic inflammation.