where X0 is the original amount of target mRNA, R0 is the original amount of b-actin mRNA, CtX is the CT value for the target and CTR is the CT value for b-actin. The CT values refer to the number of PCR cycles Sutezolid performed for each PCR product in a sample at a specific time point. Four female and eight male patient samples were used. The resulting values were examined using Grubb’s test and outliers were removed. Immunohistochemistry. Each experiment was repeated two to three times to ensure reproducibility. 10725256 Qualitative assessment of the immunohistochemical staining was evaluated by two human readers, blinded to the study details and with extensive expertise in the field. Comparison of immunoreactivity in fresh and organ cultured tissue was always performed within tissue from the same patient. Five female 27326330 and six male samples were evaluated in each group. Additionally, measurement of the fluorescence intensity in the smooth muscle cell layer of each artery section was performed in a blinded manner using the Image J software. This was performed to further evaluate the immunoreactivity and, in addition, strengthen the qualitative assessment made by the human reader. The mean 
intensities of male and female arteries incubated for 0 h and 48 h of organ culture are presented. Because of the comparative studies, immunoreactivity to individual receptors was visualized with the same microscope settings during the same day. there was no acetylcholine receptor-mediated relaxation in either male or female cerebral arteries. Furthermore, a few experiments were performed with continuous blockade of prostaglandin and NO synthesis by indomethacin and L-NAME which had no effect on the vasoconstrictor responses. Together, these results indicate a lack of functional endothelium in cultured arteries. Contractile responses to 5-carboxamidotryptamine. 5HT1B receptor-mediated contraction was studied by cumulative application of increasing concentrations of the agonist 5-CT. 5-CT induced contractile responses that followed a monophasic concentration-response curve. No significant differences between male and female cerebral arteries were observed in 5-HT1B receptor-mediated contraction. Contractile responses to angiotensin II. Ang II induced a concentration-dependent contraction at lower concentrations and dilatation at higher concentrations. These responses occurred in both male and female arteries after organ culture, but the male arteries were more sensitive to Ang II resulting in concentration-response curves that were significantly different. pEC50 for Ang II-mediated contraction was higher for males compared to females, as shown by a concentration-response curve further to the left for males. The maximum contraction to Ang II was not different between male and female arteries. Contractile responses to endothelin-1. In male cerebral arteries exposed to organ culture, application of ET-1 elicited a biphasic concentration-dependent response indicating the presence of both ETB receptors and ETA receptors. However, no ETB receptor-mediated contraction was observed in three of the male patient samples. In all female arteries, ET-1 elicited a monophasic response demonstrating presence of ETA receptors, but weak or no ETB receptor-mediated contraction. This was shown by a difference in the maximum contraction of the first phase in the ET-1 contraction and statistically different curves. ETA receptor-mediated contraction was not significantly different in male and
