Microarray and functional cluster analysis implicates transforming growth factor beta1 in endothelial cell dysfunction in a swine hemorrhagic shock model.

BACKGROUND: Trauma leading to massive hemorrhage results in widespread tissue hypoxia, anaerobic metabolism, and production of inflammatory cytokines and oxidative molecules injurious to the vascular endothelium. Although trauma-related endothelial cell pathophysiology has been extensively studied, very little is known regarding gene transcriptional changes that occur during the event, particularly in endothelia. Thus, we employed fluorescent microarray analysis of gene transcription to elucidate critical pathways and gene products involved in endothelial dysfunction. MATERIALS AND METHODS: A trauma-hemorrhage/shock (T-H/S) model mimicking the physiologic changes seen in human trauma was performed on 10 Yorkshire swine, consisting of 35% blood volume hemorrhage followed by 6 h of full resuscitation. Aortic endothelium was analyzed by microarray and functional clusters were identified through the use of Database for Annotation, Visualization, and Integrated Discovery (DAVID) software. RESULTS: Injured swine developed profound acidosis, coagulopathy, massive resuscitative fluid requirements, and microscopic changes of ischemia/reperfusion injury. While 1007 transcripts were down-regulated, 529 transcripts were up-regulated. DAVID functional clustering analysis revealed 21 significantly altered biological processes that were grouped into 12 distinct functional categories. The transforming growth factor beta (TGFß) family of genes was the most interrelated. In addition, vascular endothelial growth factor (VEGF) signaling members and leukocyte chemoattractants were also altered. CONCLUSIONS: Our model identified two major signaling pathways, TGFß and VEGF, which undergo early transcriptional changes in injured endothelial cells. Our results suggest that TGFß and VEGF may play a crucial role in the development of endothelial cell injury leading to increased vascular permeability during shock-trauma.

Transcriptional analysis of novel hormone receptors PGRMC1 and PGRMC2 as potential biomarkers of breast adenocarcinoma staging.

BACKGROUND: The expression of progesterone receptor membrane component 1 (PGRMC1) in breast cancer has generated interest in this recently discovered protein because of its role in tumorigenesis. However, correlations between patient age, PGRMC1 gene expression, breast cancer morphology, and breast cancer stage have not been adequately studied. Furthermore, very little is known about possible roles for other PGRMC isoforms in breast cancer, like PGRMC2. Thus, we examined the expression of PGRMC1 and PGRMC2 mRNA by relative quantitative PCR (RelqPCR) and determined whether transcript levels correlate with age, breast cancer staging, estrogen receptor alpha (ERα) status, and other morphometric features routinely used during the pathological examination of breast ductal adenocarcinomas. METHODS: Twenty-eight frozen or paraffin embedded breast cancer samples (ductal carcinoma in situ and stages I thru IV invasive ductal adenocarcinoma) and 10 control benign breast tissue samples were randomly selected and interrogated by RelqPCR to determine PGRMC1, 2, and ERα mRNA transcript levels. To control for slight variations in sample preparation, receptor transcript was normalized to the housekeeping gene phosphoglycerate kinase 1 (PGK1). Descriptive statistics and ANOVA of multiparametric datasets were used to correlate transcript levels with pathological staging parameters. RESULTS: PGRMC1 mRNA levels decreased significantly with patient age (Pearson's correlation -0.369; P=0.035), whereas PGRMC2 levels did not. Although the mean relative expression of PGRMC1 significantly decreased in stage II breast cancer compared with controls (P=0.050), it was no longer significant when age was considered a covariance (P=0.371). On the other hand, PGRMC2 mRNA transcript was significantly decreased in stage II breast cancer when compared to stage III cancer (P=0.028) in a manner independent of age (corrected model Bonferroni pair wise comparison, P=0.036). Furthermore, PGRMC2 levels positively correlated with ERα mRNA transcripts in patients with ER positive tumors (Pearson's correlation 0.503, P=0.096). CONCLUSIONS: Decreases in PGRMC1 mRNA are partially explained by increasing patient age. On the other hand, compared to stage III, PCRMC2 mRNA was significantly decreased in stage II adenocarcinoma of the breast in an age-independent manner. Additionally, PGRMC2 mRNA levels displayed a positive correlation with ERα transcripts. Thus, in addition to morphometric pathologic staging criteria, measurements of PGRMC2 mRNA may be useful for distinguishing low stage tumors from higher stages that require more aggressive clinical management, and may be a useful test when tumor ER IHC results are equivocal.