Cancer may arise from a cancer stem/progenitor cell that shares characteristics with its normal counterpart. We report the reconstitution of the original human prostate cancer specimen from epithelial cell lines (termed HPET for human prostate epithelial/hTERT) derived from this sample. These tumors can be described in terms of Gleason score, a classification not applied to any of the transgenic mouse models currently developed to mimic human disease. Immunohistochemical and Western blot analyses indicate that they do not express androgen receptor or p63, similar to that reported for prostate stem cells. These cell lines also express embryonic stem markers (Oct4, Nanog, and Sox2) as well as early progenitor cell markers (CD44 and Nestin) in vitro. Clonally derived HPET cells reconstitute the original human tumor in vivo and differentiate into the three prostate epithelial cell lineages, indicating that they arise from a common stem/progenitor cell. Serial transplantation experiments reconstitute the tumors, suggesting that a fraction of parental or clonally derived HPET cells have self-renewal potential. Thus, this model may enhance our understanding of human tumor development and provide a mechanism for studying cancer stem/progenitor cells in differentiation, tumorigenesis, preclinical testing, and the development of drug resistance. [Cancer Res 2007;67(10):4807-15]

The stem cell niche provides a regulatory microenvironment for cells as diverse as totipotent embryonic stem cells to cancer stem cells (CSCs) which exhibit stem cell-like characteristics and have the capability of regenerating the bulk of tumor cells while maintaining self-renewal potential. The transmembrane glycoprotein CD44 is a common component of the stem cell niche and exists as a standard isoform (CD44s) and a range of variant isoforms (CD44v) generated though alternative splicing. CD44 modulates signal transduction through post-translational modifications as well as interactions with hyaluronan, extracellular matrix molecules and growth factors and their cognate receptor tyrosine kinases. While the function of CD44 in hematopoietic stem cells has been studied in considerable detail, our knowledge of CD44 function in tissue-derived stem cell niches remains limited. Here we review CD44s and CD44v in both hematopoietic and tissue-derived stem cell niches, focusing on their roles in regulating stem cell behavior including self-renewal and differentiation in addition to cell-matrix interactions and signal transduction during cell migration and tumor progression. Determining the role of CD44 and CD44v in normal stem cell, CSC and (pre)metastatic niches and elucidating their unique functions could provide tools and therapeutic strategies for treating diseases as diverse as fibrosis during injury repair to cancer progression.

Inertial microfluidics has been attracting considerable interest in recent years due to immensely promising applications in cell separations and sorting. Despite the intense attention, the moderate efficiencies and low purity of the reported devices have hindered their widespread acceptance. In this work, we report on a simple inertial microfluidic system with high efficiency (>99%) and purity (>90%). Our system builds on the concept of two-stage inertial migration which permits precise prediction of particle or cell position within the microchannel. Our design manipulates the inertial equilibrium positions by modulating channel aspect ratio to achieve a complete separation. Here, we successfully demonstrate a complete separation of particles and isolation of rare cells in blood spiked with human prostate epithelial tumor (HPET) cells. Based on the planar structure, large separation spacing and predictable focusing, we envision promising applications and easy integration of our system with existing lab-on-a-chip systems for cell separations.

Human 15S-lipoxygenase-2 (15-LOX-2) is a recently identified lipoxygenase that has approximately 40% sequence identity to the known human 5S-, 12S-, and 15S-lipoxygenases. 15-LOX-2 has a limited tissue dis-Arachidonic acid (AA) metabolites are important mediators of a variety of physiological processes and inflammatory reactions. In addition, alterations in AA metabolism may potentially mediate key steps in certain neoplastic processes. 1-3 AA is metabolized via cyclooxygenase to prostaglandins, prostacyclin, and thromboxane, 4 and via lipoxygenases (LOX) to hydroxyeicosatetraenoic acids (HETEs) or leukotrienes (5-LOX pathway). 5,6 Until recently, three lipoxygenases were recognized in humans: a 5S-LOX found in leukocytes, a 12S-LOX found in platelets and certain epithelia, and a 15S-LOX in reticulocytes, eosinophils, macrophages, and skin. 7 Recently, in studying lipoxygenase expression in human skin, Brash et al 8 discovered a second 15S-lipoxygenase (herein referred to as 15-LOX-2). The cDNA-derived amino acid sequence of 15-LOX-2 showed only 44% identity to 5-LOX and 38% to 39% identity to 12-LOX and the reticulocyte type of 15-LOX

Inertial microfluidics has been attracting considerable interest for size-based separation of particles and cells. The inertial forces can be manipulated by expanding the microchannel geometry, leading to formation of microvortices which selectively isolate and trap particles or cells from a mixture. In this work, we aim to enhance our understanding of particle trapping in such microvortices by developing a model of selective particle trapping. Design and operational parameters including flow conditions, size of the trapping region, and target particle concentration are explored to elucidate their influence on trapping behavior. Our results show that the size dependence of trapping is characterized by a threshold Reynolds number, which governs the selective entry of particles into microvortices from the main flow. We show that concentration enhancement on the order of 100,000× and isolation of targets at concentrations in the 1/mL is possible. Ultimately, the insights gained from our systematic investigation suggest optimization solutions that enhance device performance (efficiency, size selectivity, and yield) and are applicable to selective isolation and trapping of large rare cells as well as other applications.

cAMP-regulated transcription of the human vasoactive intestinal peptide gene is dependent upon a 17-basepair DNA element located 70 base pairs upstream from the transcriptional initiation site. This element is similar to sequences in other genes known to be regulated by cAMP and to sequences in several viral enhancers. We have demonstrated that the vasoactive intestinal peptide regulatory element is an enhancer that depends upon the integrity of two CGTCA sequence motifs for biological activity. Mutations in either of the CGTCA motifs diminish the ability of the element to respond to cAMP. Enhancers containing the CGTCA motif from the somatostatin and adenovirus genes compete for binding of nuclear proteins from C6 glioma and PC12 cells to the vasoactive intestinal peptide enhancer, suggesting that CGTCA-containing enhancers interact with similar transacting factors.DNA control elements that activate transcription in response to intracellular effectors such as cAMP or products of phosphoinositol metabolism have been identified in several cellular and viral genes (1-6). These genetic elements may be particularly important in neuroendocrine cells where the activation of intracellular second messenger pathways may couple secretion and biosynthesis of neuropeptides (7,8). The genetic element responsible for cAMP-regulated transcription of the human vasoactive intestinal peptide (VIP) gene consists ofa 17-base-pair (bp) sequence located between 70 and 86 bp upstream from the transcriptional initiation site (5). This sequence in the VIP gene is similar to the cAMPresponsive elements (CREs) of the rat somatostatin, human proenkephalin, human a-chorionic gonadotrophin, and rat phosphoenolpyruvate carboxykinase genes (1-5). All of these CREs contain one or multiple copies of the conserved sequence motif CGTCA and several have been shown to have properties of enhancers (2-4). The CGTCA motif is also found in several viral enhancers such as the 18-bp repeat of the cytomegalovirus gene and the ElA-inducible element of the adenovirus E4 promoter (9, 10). The presence of the CGTCA motif in these viral enhancers suggests that similar nuclear factors may activate transcription of both viral and cellular genes. In the VIP CRE, the two CGTCA motifs are present as inverted repeats, a structure characteristic of some other enhancers (11-13). It is not known whether the palindromic structure of the VIP CRE is necessary for biological activity.In this study, three major issues were addressed. First, we sought to determine whether the VIP CRE could be categorized as an enhancer. Second, we examined the importance of the CGTCA motifs for activity of the DNA element.

Increases in neuroendocrine (NE) cells and their secretory products are closely correlated with tumor progression and androgen-independent prostate cancer. However, the mechanisms by which NE cells influence prostate cancer growth and progression, especially after androgen ablation therapy, are poorly understood. To investigate the role of NE cells on prostate cancer growth, LNCaP xenograft tumors were implanted into nude mice. After the LNCaP tumors were established, the NE mouse prostate allograft (NE-10) was implanted on the opposite flank of these nude mice to test whether NE tumor-derived systemic factors can influence LNCaP growth. Mice bearing LNCaP tumors with or without NE allografts were castrated 2 weeks after NE tumor inoculation, and changes in LNCaP tumor growth rate and gene expression were investigated. After castration, LNCaP tumor growth decreased in mice bearing LNCaP tumors alone, and this was accompanied by a loss of nuclear androgen receptor (AR) localization. In contrast, in castrated mice bearing both LNCaP and NE-10 tumors, LNCaP tumors continued to grow, had increased levels of nuclear AR, and secreted prostate-specific antigen. Therefore, in the absence of testicular androgens, NE secretions were sufficient to maintain LNCaP cell growth and androgen-regulated gene expression in vivo. Furthermore, in vitro experiments showed that NE secretions combined with low levels of androgens activated the AR, an effect that was blocked by the antiandrogen bicalutamide. Because an increase in AR level has been reported to be sufficient to account for hormone refractory prostate cancers, the NE cell population ability to increase AR level/activity can be another mechanism that allows prostate cancer to escape androgen ablation therapy.

In search for the molecular mechanisms underlying the formation of the major dense line in peripheral nerve myelin we investigated mice deficient in the myelin proteins P0 and MBP. In mice lacking both molecules axons were enwrapped by myelin-like processes devoid of the major dense line, while mice deficient in either protein showed, respectively, partial and normal compaction. Mice heterozygous for P0 but devoid of MBP showed myelin of reduced thickness around axons of normal caliber. Both molecules thus contribute to the formation of the major dense line and to the determination of myelin thickness. Furthermore, our observations modify the view that axon caliber is dependent on normal myelin.