Endometrial-peritoneal interactions during endometriotic lesion establishment.

The pathophysiology of endometriosis remains unclear but involves a complex interaction between ectopic endometrium and host peritoneal tissues. We hypothesized that disruption of this interaction would suppress endometriotic lesion formation. We hoped to delineate the molecular and cellular dialogue between ectopic human endometrium and peritoneal tissues in nude mice as a first step toward testing this hypothesis. Human endometrium was xenografted into nude mice, and the resulting lesions were analyzed using microarrays. A novel technique was developed that unambiguously determined whether RNA transcripts identified via microarray analyses originated from human cells (endometrium) or mouse cells (mesothelium). Four key pathways (ubiquitin/proteasome, inflammation, tissue remodeling/repair, and ras-mediated oncogenesis) were revealed, demonstrating communication between host mesothelial cells and ectopic endometrium. Morphometric analysis of nude mouse lesions confirmed that necrosis, inflammation, healing and repair, and cell proliferation occurred during xenograft development. These processes were entirely consistent with the molecular networks revealed by the microarray data. The transcripts detected in the xenografts overlapped with differentially expressed transcripts in a comparison between paired eutopic and ectopic endometria from human endometriotic patients. For the first time, components of the interaction between ectopic endometrium and peritoneal stromal tissues are revealed. Targeted disruption of this dialogue is likely to inhibit endometriotic tissue formation and may prove to be an effective therapeutic strategy for endometriosis.

Identification of novel VHL target genes and relationship to hypoxic response pathways.

Upregulation of hypoxia-inducible factors HIF-1 and HIF-2 is frequent in human cancers and may result from tissue hypoxia or genetic mechanisms, in particular the inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene (TSG). Tumours with VHL inactivation are highly vascular, but it is unclear to what extent HIF-dependent and HIF-independent mechanisms account for pVHL tumour suppressor activity. As the identification of novel pVHL targets might provide insights into pVHL tumour suppressor activity, we performed gene expression microarray analysis in VHL-wild-type and VHL-null renal cell carcinoma (RCC) cell lines. We identified 30 differentially regulated pVHL targets (26 of which were 'novel') and the results of microarray analysis were confirmed in all 11 novel targets further analysed by real-time RT-PCR or Western blotting. Furthermore, nine of 11 targets were dysregulated in the majority of a series of primary clear cell RCC with VHL inactivation. Three of the nine targets had been identified previously as candidate TSGs (DOC-2/DAB2, CDKN1C and SPARC) and all were upregulated by wild-type pVHL. The significance for pVHL function of two further genes upregulated by wild-type pVHL was initially unclear, but re-expression of GNG4 (G protein gamma-4 subunit/guanine nucleotide-binding protein-4) and MLC2 (myosin light chain) in a RCC cell line suppressed tumour cell growth. pVHL regulation of CDKN1C, SPARC and GNG4 was not mimicked by hypoxia, whereas for six of 11 novel targets analysed (including DOC-2/DAB2 and MLC2) the effects of pVHL inactivation and hypoxia were similar. For GPR56 there was evidence of a tissue-specific hypoxia response. Such a phenomenon might, in part, explain organ-specific tumorigenesis in VHL disease. These provide insights into mechanisms of pVHL tumour suppressor function and identify novel hypoxia-responsive targets that might be implicated in tumorigenesis in both VHL disease and in other cancers with HIF upregulation.

The psoriatic transcriptome closely resembles that induced by interleukin-1 in cultured keratinocytes: dominance of innate immune responses in psoriasis.

Psoriasis has been considered an autoimmune, T cell-mediated disorder in which adaptive immune responses predominate over those of non-antigen-specific innate immunity. To test this hypothesis, we profiled the transcriptome of psoriatic tissue and compared the data with that from cultured human keratinocytes exposed to the proinflammatory cytokine interleukin (IL)-1alpha and the Th1 cytokine interferon-gamma. When compared with patient-matched, nonlesional skin biopsies, psoriatic samples exhibited regulation of 90 transcripts including several members of the epidermal differentiation complex, molecules with antimicrobial activity, and hyperproliferation-associated keratins. Stimulation of keratinocytes with interferon-gamma resulted in regulation of 252 transcripts, with particularly strong expression of the CXCR3-binding ligands CXCL9, -10, and -11 and class II major histocompatibility complex genes, primarily those of the HLA-DR and -DP families. In contrast, the transcriptome resulting from exposure of keratinocytes to IL-1alpha elicited differences in just 19 transcripts, particularly genes within the epidermal differentiation complex and antimicrobial molecules, including PI3 and DEFB4. Major differences between the two keratinocyte transcriptomes were exhibited with only five induced IL-1alpha transcripts also regulated in the interferon-gamma set. Unexpectedly, there was a high correlation between psoriatic lesional tissue and the IL-1alpha transcriptome. These findings suggest that the inflammatory milieu in the epidermal microenvironment in psoriasis is more likely dependent on evolutionarily ancient cytokines such as IL-1, rather than those of the adaptive immune response.

Transcript profiling of functionally related groups of genes during conditional differentiation of a mammalian cochlear hair cell line.

We have used Affymetrix high-density gene arrays to generate a temporal profile of gene expression during differentiation of UB/OC-1, a conditionally immortal cell line derived from the mouse cochlea. Gene expression was assessed daily for 14 days under differentiating conditions. The experiment was replicated in two separate populations of cells. Profiles for selected genes were correlated with those obtained by RT-PCR, TaqMan analysis, immunoblotting, and immunofluorescence. The results suggest that UB/OC-1 is derived from a population of nonsensory epithelial cells in the greater epithelial ridge that have the potential to differentiate into a hair-cell-like phenotype, without the intervention of Math1. Elements of the Notch signaling cascade were identified, including the receptor Notch3, with a transient up-regulation that suggests a role in hair cell differentiation. Several genes showed a profile similar to Notch3, including the transcriptional co-repressor Groucho1. UB/OC-1 also expressed Me1, a putative partner of Math1 that may confer competence to differentiate into hair cells. Cluster analysis revealed expression profiles for neural guidance genes associated with Gata3. The temporal dimension of this analysis provides a powerful tool to study genetic mechanisms that underlie the conversion of nonsensory epithelial cells into hair cells.