A novel immunohistochemical method to estimate cell-cycle phase distribution in archival tissue: implications for the prediction of outcome in colorectal cancer

An immunohistochemical method for assessing cell-cycle phase distribution in colorectal resection specimens would enable phase data to be incorporated into diagnostic algorithms for the estimation of prognosis and response to adjuvant chemotherapy in colorectal cancer. In contrast to flow cytometry, an immunohistochemical method would also allow the phase distribution to be examined within morphologically heterogeneous regions of neoplasms. Paraffin sections of normal colon (n = 25), colonic adenoma (n = 15), and colonic adenocarcinoma (n = 30) were analysed by immunohistochemistry using antibodies against markers of cell-cycle entry, Mcm-2 and Ki67, and putative markers of the cell-cycle phase, cyclins D1 and E (putative markers of G1 phase), cyclin A (S phase), cytoplasmic cyclin B1 (G2 phase), and phosphohistone H3 (M phase). The phase specificity of each marker was assessed by examining the degree of co-expression of adjacent phase markers using double-antibody fluorescence confocal microscopy and by comparison with flow cytometric analysis performed on adjacent tissue sections. The S-phase specificity of detectable cyclin A was also assessed in combination with in situ DNA replication using fluorescence confocal microscopy. All cells expressing phase markers co-expressed Mcm-2. Adjacent phase markers were not significantly co-expressed, confirming the relative specificity of these markers in tissue sections of colon. Cell-cycle phase distribution, calculated by immunohistochemistry, compared well with phase analyses obtained by flow cytometry. No cells expressed cyclin A in the absence of active DNA replication. The S-phase labelling index, as defined by detectable cyclin A expression, showed a positive correlation with the Mcm-2 labelling index and increased in the progression from normal colon to adenocarcinoma. In conclusion, a combination of these cell-cycle phase markers can be used to calculate the distribution of cells throughout each phase of the cell cycle in colorectal tissue sections. Detectable cyclin A can be used as a surrogate marker of S phase and may be of value in predicting prognosis and response to adjuvant therapy.     

Effects of FiO2 on O2 consumption and cardiovascular and hormonal responses to hemorrhage in the goat

This study was conducted to determine whether 100% FiO2 during hemorrhage would improve maintenance of mean arterial blood pressure and to assess the cardiovascular, arginine vasopressin, and renin-angiotensin system roles in the response. This also allowed evaluation of FiO2 effects on the baroreceptor control of these hormone systems. Six conscious female goats were hemorrhaged (0.5 mL x kg(-1) x min(-1)) for 30 minutes, while breathing 11%, 21%, or 100% O2. Mean arterial blood pressure was maintained only when the goats breathed 100% O2. FiO2 did not affect O2 consumption or delivery during hemorrhage. Increases in renin and arginine vasopressin were equal or reduced with 100% FiO2 and therefore did not contribute to the improved mean arterial blood pressure maintenance. Inspiration of 100% FiO2 during hemorrhage, therefore, may improve perfusion pressure with no changes in O2 consumption or delivery. Also, 100% FiO2 eliminated the negative correlation between both hormones and right atrial pressure, suggesting altered baroreceptor control of the hormones.