The RNA binding protein HuR differentially regulates unique subsets of mRNAs in estrogen receptor negative and estrogen receptor positive breast cancer

<p>Abstract</p> <p>Background</p> <p>The discordance between steady-state levels of mRNAs and protein has been attributed to posttranscriptional control mechanisms affecting mRNA stability and translation. Traditional methods of genome wide microarray analysis, profiling steady-state levels of mRNA, may miss important mRNA targets owing to significant posttranscriptional gene regulation by RNA binding proteins (RBPs).</p> <p>Methods</p> <p>The ribonomic approach, utilizing RNA immunoprecipitation hybridized to microarray (RIP-Chip), provides global identification of putative endogenous mRNA targets of different RBPs. HuR is an RBP that binds to the AU-rich elements (ARE) of labile mRNAs, such as proto-oncogenes, facilitating their translation into protein. HuR has been shown to play a role in cancer progression and elevated levels of cytoplasmic HuR directly correlate with increased invasiveness and poor prognosis for many cancers, including those of the breast. HuR has been described to control genes in several of the acquired capabilities of cancer and has been hypothesized to be a tumor-maintenance gene, allowing for cancers to proliferate once they are established.</p> <p>Results</p> <p>We used HuR RIP-Chip as a comprehensive and systematic method to survey breast cancer target genes in both MCF-7 (estrogen receptor positive, ER+) and MDA-MB-231 (estrogen receptor negative, ER-) breast cancer cell lines. We identified unique subsets of HuR-associated mRNAs found individually or in both cell types. Two novel HuR targets, <it>CD9 </it>and <it>CALM2 </it>mRNAs, were identified and validated by quantitative RT-PCR and biotin pull-down analysis.</p> <p>Conclusion</p> <p>This is the first report of a side-by-side genome-wide comparison of HuR-associated targets in wild type ER+ and ER- breast cancer. We found distinct, differentially expressed subsets of cancer related genes in ER+ and ER- breast cancer cell lines, and noted that the differential regulation of two cancer-related genes by HuR was contingent upon the cellular environment.</p

Hepatic damage during acute pancreatitis in the rat

We studied the alterations in the metabolism of liver mitochondria in rats with acute pancreatitis. Male Wistar rats were allocated to a control group (group I) and to five other groups corresponding to 2, 4, 12, 24 and 48 h after the induction of acute pancreatitis by the injection of 5% sodium taurocholate into the pancreatic duct. Sham-operated animals were submitted to the same surgical steps except for the induction of acute pancreatitis. Mitochondrial oxidation and phosphorylation were measured polarographically by determining oxygen consumption without ADP (basal respiration, state 4) and in the presence of ADP (activated respiration, state 3). Serum amylase, transaminases (ALT and AST) and protein were also determined. Ascitic fluid, contents of amylase, trypsin and total protein were also determined and arterial blood pressure was measured in all groups. In ascitic fluid, trypsin and amylase increased reaching a maximum at 2 and 4 h, respectively. Serum amylase increased at 2 h reaching a maximum at 4 h. Serum transaminase levels increased at 12 and 24 h. After 2 h (and also 4 h) there was an increase in state 4 respiration (45.65 ± 1.79 vs 28.96 ± 1.50) and a decrease in respiration control rate (3.53 ± 0.09 vs 4.45 ± 0.08) and in the ADP/O ratio (1.77 ± 0.02 vs 1.91 ± 0.01) compared to controls (P<0.05). These results indicate a disruption of mitochondrial function, which recovered after 12 h. In the 48-h groups there was mitochondrial damage similar to that occurring in ischemic lesion. Beat-to-beat analysis (30 min) showed that arterial blood pressure remained normal up to 24 h (111 ± 3 mmHg) while a significant decrease occurred in the 48-h group (91 ± 4 mmHg). These data suggest biphasic damage in mitochondrial function in acute pancreatitis: an initial uncoupled phase, possibly secondary to enzyme activity, followed by a temporary recovery and then a late and final dysfunction, associated with arterial hypotension, possibly related to ischemic damage